Peptide for vaccine

ABSTRACT

The present invention relates to compositions comprising peptides for preventing or treating allergy to house dust mites, and in particular to optimal combinations of peptides for preventing or treating said allergy.

RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.14/150,045 (allowed), filed on Jan. 8, 2014, which is a continuation ofU.S. Pat. No. 8,652,485, issued on Feb. 18, 2014, which is a nationalstage filing under 35 U.S.C. 371 of International Patent Application No.PCT/GB2008/002780, filed Aug. 15, 2008, which claims the benefit ofBritish Patent Application No. 0715949.4, filed Aug. 15, 2007, BritishPatent Application No. 0716224.1, filed Aug. 20, 2007, and BritishPriority Application No. 0723337.2, filed Nov. 28, 2007.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has beensubmitted in ASCII format via EFS-Web, and is hereby incorporated byreference in its entirety. Said ASCII copy, created on Oct. 17, 2016, isnamed JKJ-010USCN2_Sequence_Listing.txt and is 232,786 bytes in size.

FIELD OF THE INVENTION

The present invention relates to compositions comprising peptides forpreventing or treating allergy to house dust mites, and in particular tooptimal combinations of peptides for preventing or treating saidallergy.

BACKGROUND OF THE INVENTION

T-cell antigen recognition requires antigen presenting cells (APCs) topresent antigen fragments (peptides) on their cell surface inassociation with molecules of the major histocompatibility complex(MHC). T cells use their antigen specific T-cell receptors (TCRs) torecognise the antigen fragments presented by the APC. Such recognitionacts as a trigger to the immune system to generate a range of responsesto eradicate the antigen which has been recognised.

Recognition of external antigens by the immune system of an organism,such as man, can in some cases result in diseases, known as atopicconditions. Examples of the latter are the allergic diseases includingasthma, atopic dermatitis and allergic rhinitis. In this group ofdiseases, B lymphocytes generate antibodies of the IgE class (in humans)which bind externally derived antigens, which are referred to in thiscontext as allergens since these molecules elicit an allergic response.Production of allergen-specific IgE is dependent upon T lymphocyteswhich are also activated by (are specific for) the allergen.Allergen-specific IgE antibodies bind to the surface of cells such asbasophils and mast cells by virtue of the expression by these cells ofsurface receptors for IgE.

Crosslinking of surface bound IgE molecules by allergen results indegranulation of these effector cells causing release of inflammatorymediators such as histamine, 5-hydroxtryptamine and lipid mediators suchas the sulphidoleukotrienes. In addition to IgE-dependent events,certain allergic diseases such as asthma are characterised byIgE-independent events.

Allergic IgE-mediated diseases are currently treated with agents whichprovide symptomatic relief or prevention. Examples of such agents areanti-histamines, P2 agonists, and glucocorticosteroids. In addition,some IgE-mediated diseases are treated by desensitisation proceduresthat involve the periodic injection of allergen components or extracts.Desensitisation treatments may induce an IgG response that competes withIgE for allergen, or they may induce specific suppressor T cells thatblock the synthesis of IgE directed against allergen. This form oftreatment is not always effective and poses the risk of provokingserious side effects, particularly general anaphylactic shock. This canbe fatal unless recognised immediately and treated with adrenaline. Atherapeutic treatment that would decrease or eliminate the unwantedallergic-immune response to a particular allergen, without altering theimmune reactivity to other foreign antigens or triggering an allergicresponse itself would be of great benefit to allergic individuals.

House dust mites are universally recognised as a major cause of allergicdiseases in humans and animals, including asthma, allergic rhinitis andallergic dermatitis. Two closely related species of mite are responsiblefor the majority of house dust mite allergy worldwide. These areDermatophagoides pteronyssinus (predominantly in Europe) andDermatophagoides farinae (predominantly in America). House dust miteallergens are mainly derived from proteins from the lining of the mitegut, which are present in the faeces, and are typically referred to asDer p (for D. pteronyssinus) or Der f (for D. farinae) proteins. Anaverage mite will produce approximately 20 faecal pellets each day ofits life: twice its own body weight. One gram of dust can typicallycontain up to 500 mites, while a mattress can hold more than twomillion. The amount of mite material present increases with age. Onetenth of the weight of a six-year old pillow can consist of mites andmite debris. In a carpet, there will typically be between 1,000 and10,000 mites per square meter.

Allergic diseases, particularly asthma, are a huge and expanding problemin the industrialised nations of the world. It has been calculated that5-10% of the population of the major industrialised nations suffers fromasthma. Of those, approximately one fifth will have severe asthmarequiring frequent hospitalisation. The cost of asthma within the UnitedStates has been calculated as $12.6 billion (£7.9 billion) per year.Figures for Europe are even higher. A Canadian study estimated the costsof asthma as averaging £21 per year for every member of the populationof the major industrialised nations. 2,000 people every year will die asa result of asthma in the United Kingdom alone.

Asthma is a chronic disease caused by allergic reactions and irritationwithin the respiratory system. Between 50% and 90% of asthmatics whoreact to airborne material are sensitive to dust mite allergens, and inone British study 10% of the general population reacted to dust miteallergens. Almost two hundred million Americans live in areas severelyaffected by house dust mite infestation. Sensitisation to this materialoccurs in childhood, mainly between three and six months of age butasthma is lifelong.

A therapeutic or preventative treatment would therefore be of greatbenefit to humans that suffer or are at risk of suffering from housedust mite allergy.

SUMMARY OF THE INVENTION

The present inventors have discovered that certain combinations ofpeptide fragments derived from the Group 1 dust mite allergen (Der p 1,Der f 1), Group 2 dust mite allergen (Der p 2, Der f 2) and Group 3 dustmite allergen (Der p 7, Der f 7) are particularly useful indesensitising individuals to these allergens. The polypeptidecombinations of the invention have been selected for their ability toinduce a cytokine response in a high proportion of subjects from a panelof house dust mite allergic individuals.

The polypeptides of the invention were initially selected as T cellepitopes through use of both in silico and in vitro assessments ofpeptide-MHC binding characteristics. See for example Table 3 whichdemonstrates the ability of a range of peptides derived from the aboveallergens to bind to multiple DR types in MHC class II binding assays.Additional epitopes were identified by homology. These candidatepolypeptides were then further screened for potential use intolerisation.

A difficulty associated with approaches to desensitisation based onpeptide immunisation lies in how to select an appropriate size andregion of the allergen as the basis for the peptide to be used forimmunisation. The size of the peptide of choice is crucial. If thepeptide is too small, the vaccine would not be effective in inducing animmunological response. If the peptides are too large, or if the wholeantigen is introduced into an individual, there is the risk of inducingadverse reactions, such as anaphylaxis, which may be fatal.

The polypeptides of the invention have been selected to retain T cellspecificity whilst being small enough in size to not possess significanttertiary structure that would enable them to retain the conformation ofan IgE-binding epitope of the whole molecule. The polypeptides of theinvention therefore do not induce significant crosslinking of adjacentspecific IgE molecules on cells such as mast cells and basophils andconsequently do not cause significant histamine release.

An advantage of the invention is the ability of the peptides to broadlytarget Major Histocompatibility Complex (MHC) molecules. T cellreceptors (TCRs) are highly variable in their specificity. Variabilityis generated, as with antibody molecules, through gene recombinationevents within the cell. TCRs recognise antigen in the form of shortpeptides bound to molecules encoded by the genes of the MajorHistocompatibility Complex (MHC). These gene products are the samemolecules that give rise to “tissue types” used in transplantation andare also referred to as Human Leukocyte Antigen molecules (HLAs) whichterms may be used interchangeably. Individual MHC molecules possesspeptide binding grooves which, due to their shape and charge are onlycapable of binding a limited group of peptides. The peptides bound byone MHC molecule may not necessarily be bound by other MHC molecules.

When a protein molecule such as an antigen or allergen is taken up byantigen presenting cells such as B lymphocytes, dendritic cells,monocytes and macrophages, the molecule is enzymatically degraded withinthe cell. The process of degradation gives rise to peptide fragments ofthe molecule which, if they are of the appropriate size, charge andshape, may then bind within the peptide binding groove of certain MHCmolecules and be subsequently displayed upon the surface of antigenpresenting cells. If the peptide/MHC complexes are present upon theantigen presenting cell surface in sufficient numbers they may thenactivate T cells which bear the appropriate peptide/MHC-specific T cellreceptors.

Due to the polymorphic nature of the MHC, individuals in an outbredpopulation such as man will express different combinations of MHCmolecules on their cell surfaces. Since different MHC molecules can binddifferent peptides from the same molecule based on the size, charge andshape of the peptide, different individuals will display a differentrepertoire of peptides bound to their MHC molecules. Identification ofuniversal MHC-binding peptide epitopes in an outbred population such asman is more difficult than in inbred animals (such as certain strains oflaboratory mice). On the basis of differential MHC expression betweenindividuals and the inherent differences in peptide binding andpresentation which this brings, it is unlikely that a single peptide canbe identified which will be of use for desensitisation therapy in man.

The peptide combinations of the invention, however, provide a broadcoverage of efficacy over the human population by targeting multipledifferent MHC molecules. A vaccine formulated with the peptides of theinvention would therefore have broad utility.

The inventors' work has produced peptide combinations with the followingcharacteristics:

-   -   the combination induces a cytokine response in a high proportion        of subjects from a panel of house dust mite allergic individuals    -   the peptides of the combinations are soluble.

Accordingly, the present invention provides a composition for use inpreventing or treating allergy to house dust mites by tolerisationcomprising at least one polypeptide selected from HDM203B (SEQ ID 83),HDM201 (SEQ ID 80), HDM205 (SEQ ID 85), HDM203A (SEQ ID 82), HDM202 (SEQID 81), SEQ ID NO's 1 to 79, 84, or 86 to 104 (that is any one of SEQ IDNO's. 1 to 104) or a variant thereof. Typically, the compositioncomprises at least four polypeptides, wherein the polypeptides areindependently selected from any of the following:

(i) a polypeptide of SEQ ID NO's 1 to 104; or

(ii) a variant of a polypeptide according to (i), wherein said variantis a polypeptide of length 9 to 30 amino acids that comprises a regionconsisting of:

-   -   any of the sequences of (i); or    -   a sequence which has at least 65% homology to any of the        sequences of (i) which sequence is capable of tolerising an        individual to any of the sequences of (i); or

(iii) a variant of a polypeptide according to (i), wherein said variantis a polypeptide of length 9 to 30 amino acids that comprises a regionconsisting of a sequence that represents either:

-   -   a fragment of any of the sequences of (i); or    -   a homologue of a fragment of any of the sequences of (i), which        sequence is capable of tolerising an individual to any of the        sequences of (i) and has a length of at least 9 amino acids, and        wherein said homologue has at least 65% homology to any 9        contiguous amino acids in any of the sequences of (i).

DESCRIPTION OF THE DRAWINGS

FIG. 1—Sequence comparison of Der p 1 (Residues 17-320 of SEQ ID NO:143) versus Der f 1 (Residues 17-321 of SEQ ID NO: 151) (FIG. 1A), Der p2 (SEQ ID NO: 144) versus Der f 2 (SEQ ID NO: 152) (FIG. 1B) and Der p 7(Residues 1-213 of SEQ ID NO: 149) versus Der f 7 (Residues 1-213 of SEQID NO: 155) (FIG. 1C). Regions containing epitopes are highlighted ingrey. Locations of specific peptides of the invention are indicated bylines above or below the sequence. The sequence of Der p 1 is thepublically available sequence with NCBI Accession No. P08176. Thecorresponding sequences for Der p 2 and Der p 7 (Table 6) are NCBIAccession Nos. P49278 and P49273, respectively. The sequence for Der f 1is taken from NCBI Accession No. P16311, Der f 2 is from NCBI AccessionNo. Q00855 and Der f 7 is from NCBI Accession No. Q26456.

FIG. 2 shows the percentage of individuals responsive to differentpeptides of the invention measured by production of IL13 or IFN-gamma.

FIG. 3 and FIG. 4 show the percentage of individuals responsive todifferent peptide combinations of the invention measured by productionof IL13 or IFN-gamma.

DESCRIPTION OF THE SEQUENCES MENTIONED HEREIN

SEQ ID NOS: 1 to 104 provide the polypeptide sequences of the inventionas set out in Tables 3 to 8. SEQ ID NOS. 105 onwards provide additionalsequences.

DETAILED DESCRIPTION OF THE INVENTION

The invention concerns peptides and combinations of peptides which canbe used in tolerisation. Such peptides may comprise, consist of, orconsist essentially of the sequences shown in any of HDM203B (SEQ ID83), HDM201 (SEQ ID 80), HDM205 (SEQ ID 85), HDM203A (SEQ ID 82), HDM202(SEQ ID 81), SEQ ID NO's 1 to 79, 84, or 86 to 104 (that is any one ofSEQ ID NO's. 1 to 104). Variants of these specific peptides may also beused. The variants may comprise, consist of, or consist essentially ofsequences which are fragments of either any of SEQ ID NO's 1 to 104 orhomologues of any of SEQ ID NO's 1 to 104.

In one embodiment the invention relates to a composition for use inpreventing or treating allergy to house dust mites. The compositiontypically comprises or consists at least four, five, six, seven, eight,nine, ten, eleven, or twelve polypeptides, up to a maximum of thirteen.In other words, the composition comprises between four and thirteenpolypeptides. The polypeptides are independently selected from any ofthe following:

(i) a polypeptide of SEQ ID NO's 1 to 104; or

(ii) a variant of a polypeptide according to (i), wherein said variantis a polypeptide of length 9 to 30 amino acids that comprises a regionconsisting of:

-   -   any of the sequences of (i); or    -   a sequence which has at least 65% homology to any of the        sequences of (i) which sequence is capable of tolerising an        individual to any of the sequences of (i), or

(iii) a variant of a polypeptide according to (i), wherein said variantis a polypeptide of length 9 to 30 amino acids that comprises a regionconsisting of a sequence that represents either:

-   -   a fragment of any of the sequences of (i); or    -   a homologue of a fragment of any of the sequences of (i), which        sequence is capable of tolerising an individual to any of the        sequences of (i) and has a length of at least 9 amino acids, and        wherein said homologue has at least 65% homology to any 9        contiguous amino acids in any of the sequences of (i).

The invention also provides products and formulations comprising thepolypeptides of the invention and compositions, products and vectorscomprising polynucleotides capable of expressing the polypeptides of theinvention for use in preventing or treating house dust mite allergy bytolerisation. Such tolerisation will typically be to an epitope (forexample a MHC class II epitope) present in any of SEQ ID NO's 1 to 104.

Peptide Fragments of Group 1, Group 2 and Group 7 Dust Mite Allergens

The major allergens of the House dust mite include the Group 1 dust miteallergen (Der p 1, Der f 1), Group 2 dust mite allergen (Der p 2, Der f2) and Group 3 dust mite allergen (Der p 7, Der f 7), wherein Der p “X”and Der f “X” indicate that the protein “X” is a homologue deriving fromD. pteronyssinus and D. farinae respectively. As shown in FIG. 1, eachof the Der p proteins is highly homologous to its corresponding Der fprotein.

The regions comprising MHC Class II-binding T cell epitopes areparticularly highly conserved between the Der p and Der f homologues ofa given protein. Peptides derived from the relevant regions of forexample, protein 1 of either D. pteronyssinus or D. farinae aretherefore suitable for use in preventing or treating house dust miteallergy by tolerisation to the Group 1 dust mite allergen. Similarlypeptides derived from the relevant regions of protein 2 from eitherspecies are suitable for use in preventing or treating house dust miteallergy by tolerisation to the Group 2 dust mite allergen, and peptidesderived from the relevant regions of protein 7 from either species aresuitable for use in preventing or treating house dust mite allergy bytolerisation to the Group 7 dust mite allergen.

The Group 1 allergen is a cysteine protease homologous to papain. Thisenzyme has been found to cleave occludin, a protein component ofintercellular tight junctions. This reveals one possible reason for theallergenicity of certain enzymes. By destroying the integrity of thetight junctions between epithelial cells, Der p 1 and Der f 1 may gainabnormal access to subepithelial antigen-presenting cells, resident mastcells, and eosinophils.

The function of the Group 2 allergen is not known, although Der p 2 andDer f 2 show distant homology to a family of lipid-binding proteins.Serum IgE levels in response to stimulation with Der p 2 in vivo havebeen shown to represent approximately one third of the total serum IgEresponse to stimulation with whole mite extracts.

The function of the Group 7 allergen is also not known. Serum IgE levelsin response to stimulation with Der p 7 in vivo have been shown torepresent approximately one fifth of the total serum IgE response tostimulation with whole mite extracts.

The peptides of the invention are derived from the Group 1, Group 2 andGroup 3 dust mite allergens as shown in Tables 3 to 8. The terms“peptide” and “polypeptide” are used interchangeably herein. The aboveproteins are also referred to herein as “the allergens”.

Tables 3 to 8 set out the sequences of the peptides of the invention,indicating the parent protein from which each peptide derives. Thesequences in Tables 4 to 6 are arranged in pairs. In each pair the uppersequence has been selected as a T cell epitope through use ofpeptide-MHC binding assays. The lower sequence has been selected by ahomology search within the sequence of the alternative protein in thegiven dust mite allergen Group. For example, peptide HDM01 in Table 4derives from Der p 1, the homologous sequence below it derives from Derf 1.

Peptide Combinations

The composition typically comprises a combination of at least fourdifferent polypeptides of the invention, up to a maximum of thirteendifferent polypeptides. Accordingly, the composition of the inventionmay consist of four, five, six, seven, eight, nine, ten, eleven, twelveor thirteen peptides.

The composition of the invention may typically comprises at least onepolypeptide or variant thereof (for example a functional variant)selected from a peptide which derives from each of Der p 1, Der p 2 andDer p 7 (or the Der f equivalents). The polypeptide combinations in thecomposition of the invention are selected to provide as broad a coverageof the human population as possible, i.e. the composition of theinvention will produce an immune response in a high proportion of dustmite allergic individuals, preferably more than 30%, 40%, 45%, 50%, 60%or 70% of dust mite allergic individuals in a panel or population ofsuch individuals. The number of individuals in a population of dust miteallergic individuals may be any suitable number, typically at least 20,30, 40, 50, 60, 70, 80, or at least 100 individuals. Preferably thepopulation has MHC allele frequencies within the range of frequenciesthat are representative of the Caucasian population. Referencepopulation allele frequencies for 11 common DRB1 allele families areshown in Table 1 (Data from HLA Facts Book, Parham and Barber).

The composition of the invention typically comprises at least onepolypeptide selected from a polypeptide of HDM203B (SEQ ID 83), HDM202(SEQ ID 81), HDM201 (SEQ ID 80), HDM205 (SEQ ID 85), HDM203A (SEQ ID82), or a variant thereof. The composition preferably comprises at leasttwo, three or four polypeptides independently selected from apolypeptide of HDM203B (SEQ ID 83), HDM202 (SEQ ID 81), HDM201 (SEQ ID80), HDM205 (SEQ ID 85), HDM203A (SEQ ID 82), or a variant thereof, withthe proviso that no more than one polypeptide or variant of SEQ ID NOS:82 and 83 is selected.

Particular variants of HDM202 (SEQ ID 81) are HDM202D (SEQ ID 102;FKNRFLMSAEA), HDM202E (SEQ ID 103; FKNRFLMSAE) and HDM202H (SEQ ID 104;EFKNRFLMSAE), which are truncations of the HDM202 sequence. It isenvisaged that each of these sequences can be modified to add at leastone (and upto 6) residues at the N and/or C terminus selected from R, K,H, E and D.

Optionally, the composition may additionally comprise at least oneadditional polypeptide selected from a polypeptide of any of SEQ ID NOS:5, 51, 52, 100, 101, 72, 73, 74, or a variant thereof. The at least oneadditional polypeptide is preferably a polypeptide of any of SEQ ID NOS:51, 73, 100 and 101.

Optionally, the composition may additionally comprise at least oneadditional polypeptide selected from a polypeptide of any of SEQ ID NOS:1, 9, 21, 24, 48, 54, 56, 57, 62, 63, 65, 76, 84 and 86, or a variantthereof. The at least one additional polypeptide is preferably apolypeptide of any of SEQ ID NOS: 63 and 65, or a variant thereof.

More specifically, in one embodiment, the invention therefore provides acomposition comprising between four and thirteen polypeptides,consisting of:

-   -   a) at least one of the polypeptides of SEQ ID NOS. 83 and 82, or        variants thereof, preferably SEQ ID NO. 83;    -   b) at least two of the polypeptides of SEQ ID NOS. 80, 81 and        85, or variants thereof; and optionally    -   c) at least one of the polypeptides of any of SEQ ID NOS: 5, 51,        52, 100, 101, 72, 73, and 74, or a variant thereof, preferably        SEQ ID NOS: 51, 73, 100 and 104 or a variant thereof; and/or    -   d) at least one of the polypeptides of any of SEQ ID NOS: 1, 9,        21, 24, 48, 54, 56, 57, 62, 63, 65, 76, 84 and 86, or a variant        thereof, preferably SEQ ID NOS: 63 and 65, or a variant thereof.

In other words, one specific embodiment of the invention provides acomposition for use in the prevention or treatment of dust mite allergyby tolerisation comprising between four and thirteen peptide sequences,wherein the composition consists of:

-   -   a) at least one of the polypeptides with the following        sequences:

HDM203B DLRQMRTVTPIRMQGGSGS (SEQ ID NO. 83) and HDM203ADLRQMRTVTPIRMQGGCGS (SEQ ID NO. 82);

-   -   or a variant thereof, and;    -   b) at least two of the polypeptides with the following        sequences:

HDM201 ESVKYVQSNGGAI (SEQ ID NO. 80); HDM202DEFKNRFLMSAEAFE (SEQ ID NO. 81); and HDM205 SYYRYVAREQS (SEQ ID NO. 85)

-   -   or variants thereof and optionally;    -   c) at least one of the polypeptides with the following        sequences:

HDM09A REALAQTHSAIAVI (SEQ ID NO. 5); HDM03DRNQSLDLAEQELVDSASQH (SEQ ID NO. 51); HDM03E RNQSLDLAEQELVD

ASQH* (SEQ ID NO. 52); HDM03V EQELVDSASQHG (SEQ ID NO. 100); HDM03WELVDSASQHG (SEQ ID NO. 101); HDM101 NYCQIYPPNVNKIREA (SEQ ID NO. 72);HDM101A NYSQIYPPNVNKIREA (SEQ ID NO. 73); and HDM101B NY

QTYPPNVNKIREA* (SEQ ID NO. 74)

-   -   or a variant thereof, and/or;    -   d) at least one of the polypeptides with the following        sequences:

HDM01 IDLRQMRTVTPIR (SEQ ID NO. 1); HDM21AKPFQLEAVFEANQNTK (SEQ ID NO. 9); HDM48 TAIFQDTVRAEMTK (SEQ ID NO. 21);HDM51A VDFKGELAMRNIEAR (SEQ ID NO. 24); HDM01AIDLRQMRTVTPIRMQGGSG (SEQ ID NO. 48); HDM06ARYVAREQSSRRP (SEQ ID NO. 54); HDM07 PNVNKIREALAQT (SEQ ID NO. 56);HDM19A DQVDVKDSANHEIKK (SEQ ID NO. 57); HDM23CGLEVDVPGIDPNASH (SEQ ID NO. 62); HDM26B GVLASAIATHAKIR (SEQ ID NO. 63);HDM35A RGLKQMKRVGDANV (SEQ ID NO. 65); HDM102ANAQRFGISNYSQI (SEQ ID NO. 76); HDM204SAYLAYRNQSLDLA (SEQ ID NO. 84); and HDM206DNGYGYFAANIDLMMIEE (SEQ ID NO. 86) or a variant thereof. *

 = aminobutyric acid.

It will be appreciated that (a) to (d) above represent stringent andhighly selective criteria for the identification of suitablecombinations of the invention. For example, if one were to select eightpeptides at random from the sequences of the invention there would benearly 100 billion possible combinations to choose from. By contrast, itis useful to consider an example of a combination of eight polypeptidesin which the above criteria are applied. For example, consider acombination wherein the following polypeptides are selected:

i) any two of the polypeptides of SEQ ID NOS. 80, 81 and 85 and at leastone of the polypeptides of SEQ ID NOS. 82 and 83; and

ii) two further polypeptides selected from the polypeptides of any ofSEQ ID NOS: 5, 51, 52, 72, 73, 74, 100 and 101; and finally

iii) two further polypeptides selected from the polypeptides of any ofSEQ ID NOS: 1, 9, 21, 24, 48, 54, 56, 57, 62, 63, 65, 76, 84 and 86.

Based on such a selection, the number of possible combinationsrepresents less than 0.0006% of the total available combinations if thecriteria determined by the inventors are not applied.

On the basis of the above, a particularly preferred combination of theinvention comprises or consists of the polypeptides of HDM201 (SEQ ID80), HDM203B (SEQ ID 83), HDM205 (SEQ ID 85), HDM03W (SEQ ID 101),HDM101A (SEQ ID 73), HDM26B (SEQ ID 63), HDM35A (SEQ ID 65), andoptionally SEQ ID NO. 24, or variants thereof.

Another preferred combination of the invention comprises or consists ofthe polypeptides of HDM201 (SEQ ID 80), HDM203B (SEQ ID 83), HDM205 (SEQID 85) and HDM03W (SEQ ID 101).

Subject to the above, the composition may optionally comprise furtherpolypeptides up to a total of thirteen unique polypeptides. Thesefurther polypeptides relate to (i.e. are typically homologues and/orfragments of) the other sequences, i.e. SEQ ID NOS: 1 to 104, that arenot amongst the polypeptides already selected. The further peptides aretypically functional variants of one of the peptides of SEQ ID NO's 1 to104. The further polypeptides may be identical to any of SEQ ID NOS: 1to 104. The composition may therefore comprise up to thirteen differentpolypeptides as provided in any of SEQ ID NO: 1 to 104. However, theoptional further polypeptides do not need to be 100% identical to any ofSEQ ID NO: 1 to 104. They are preferably at least 65% identical to atleast 9 (for example at least 10, 11, 12 or 13) or more contiguous aminoacids in any of SEQ ID NO: 1 to 104, not already selected amongst thepreviously selected polypeptide(s). These contiguous amino acids maycomprise a MHC class II epitope, for example which binds to any of theMHC molecules mentioned herein. In other words, the composition mayoptionally comprise further polypeptides up to a total of thirteenunique polypeptides, wherein the further polypeptides:

(i) comprise a sequence having at least 65% sequence identity to atleast 9 or more contiguous amino acids in any of SEQ ID NO: 1 to 104above not selected in (a) to (d) above; and

(ii) are 9 to 30 amino acids in length.

wherein each different polypeptide is for simultaneous, separate orsequential use in the prevention or treatment of dust mite allergy bytolerisation. 1 to 104

In more detail therefore, the invention provides a product containingbetween four and thirteen polypeptides as defined in (a) to (d) above;and optionally:

-   -   (e) A polypeptide:        -   (i) comprising sequence having at least 65% sequence            identity to at least 9 or more contiguous amino acids in any            of SEQ ID NO: 1 to 104 not selected in a), to d) above; and        -   (ii) 9 to 30 amino acids in length; and optionally    -   (f) A polypeptide as defined in e), but additionally not        selected in d); and optionally    -   (g) A polypeptide as defined in e), but additionally not        selected in e) to f) above; and optionally    -   (h) A polypeptide as defined in e), but additionally not        selected in e) to g) above; and optionally    -   (i) A polypeptide as defined in e), but additionally not        selected in e) to h) above; and optionally    -   (j) A polypeptide as defined in e), but additionally not        selected in e) to i) above; and optionally    -   (k) A polypeptide as defined in e), but additionally not        selected in e) to j) above) above; and optionally    -   (l) A polypeptide as defined in e), but additionally not        selected in e) to k) above; and optionally    -   (m) A polypeptide as defined in e), but additionally not        selected in e) to 1) above; and optionally    -   (n) A polypeptide as defined in e), but additionally not        selected in e) to m) above; and optionally    -   (o) A polypeptide as defined in e), but additionally not        selected in e) to n) above; and optionally    -   (p) A polypeptide as defined in e), but additionally not        selected in e) to o) above for simultaneous, separate or        sequential use in the prevention or treatment of dust mite        allergy by tolerisation.

Another embodiment of the invention is a composition for use inpreventing or treating allergy to house dust mites by tolerisationcomprising one or more polypeptide, wherein the polypeptide is selectedfrom any of the following:

-   -   (i) a polypeptide of any of HDM203B (SEQ ID 83), HDM202 (SEQ ID        81), HDM201 (SEQ ID 80), HDM205 (SEQ ID 85), HDM203A (SEQ ID        82), SEQ ID NO's 1 to 79, 84, or 86 to 104 (that is any one of        SEQ ID NO's. 1 to 104); or    -   (ii) a variant of a polypeptide according to (i), wherein said        variant is a polypeptide of length 9 to 30 amino acids that        comprises a region consisting of:        -   any of the sequences of (i); or        -   a sequence which has at least 65% homology to any of the            sequences of (i) which sequence is capable of tolerising an            individual to any of the sequences of (i), or    -   (iii) a variant of a polypeptide according to (i), wherein said        variant is a polypeptide of length 9 to 30 amino acids that        comprises a region consisting of a sequence that represents        either:        -   a fragment of any of the sequences of (i); or        -   a homologue of a fragment of any of the sequences of (i),            which sequence is capable of tolerising an individual to any            of the sequences of (i) and has a length of at least 9 amino            acids, and wherein said homologue has at least 65% homology            to any 9 contiguous amino acids in any of the sequences of            (i).

The compositions or products of the invention may comprise variants ofany of sequences defined above. The variant typically comprises 1, 2, 3or more of the MHC class II epitopes present in the correspondingpeptide of SEQ ID NO: 1 to 104.

Functional variants are mentioned herein. Such variants may be able totolerise an individual to a class II MHC epitope present in thecorresponding peptide of SEQ ID NO: 1 to 104, and thus it will typicallycomprise sequence that binds to the same MHC class II molecule and/or isrecognised by a T cell which recognises the corresponding epitope in thepolypeptide of SEQ ID NO: 1 to 104.

Variants of SEQ ID NO's 1 to 104 may be fragments derived by truncation.Truncation refers to the removal of one, two, three, four, five, six,seven, eight, nine, ten or more amino acids from the N and/or C-terminalends of a polypeptide of SEQ ID NOS. 1 to 104. Examples of suitabletruncations are provided for illustrative purposes in Example 5. Inparticular, truncations of SEQ ID NO. 81 are provided as SEQ ID NO's:102 to 104. Similarly, a number of the preferred variants of HDM03 (SEQID NOS: 89 to 101) are truncations. Particularly preferred truncationsof HDM03 are HDM03V and HDM 03W (SEQ ID 100 and 101).

Fragments may also be generated by one or more internal deletions,provided that the core 9 amino acids that makes up the T cell epitope isnot substantially disrupted.

For example, a variant of SEQ ID NO: 1 may comprise a fragment of SEQ IDNO: 1, i.e. a shorter sequence. This may include a deletion of one, two,three, four, five, six, seven, eight, nine, ten or more amino acids fromthe N-terminal end of SEQ ID NO: 1 or from the C-terminal end of SEQ IDNO: 1. Such deletions may be made from both ends of SEQ ID NO: 1. Avariant of SEQ ID NO: 1 may include additional amino acids (for examplefrom the sequence of the parent protein from which the peptide derives)extending beyond the end(s) of SEQ ID NO: 1. A variant may include acombination of the deletions and additions discussed above. For example,amino acids may be deleted from one end of SEQ ID NO: 1, but additionalamino acids from the full length parent protein sequence may be added atthe other end of SEQ ID NO: 1. The same discussion of variants abovealso applies to SEQ ID NOS: 2 to 104.

A variant peptide may include one or more amino acid substitutions fromthe amino acid sequence of any of SEQ ID NOS: 1 to 104 or a fragmentthereof. A variant peptide may comprise sequence having at least 65%sequence identity to at least 9 or more contiguous amino acids in any ofSEQ ID NOS: 1 to 104. More preferably a suitable variant may comprise atleast 70%, at least 75%, at least 80%, at least 85%, at least 90%, atleast 95%, or at least 98% amino acid identity to at least 9 contiguousamino acids of any of SEQ ID NO: 1 to 104. This level of amino acididentity may be seen at any section of the peptide, although it ispreferably the core region. The level of amino acid identity is over atleast 9 contiguous amino acids but it may be at least 10, 11, 12, 13,14, 15 or at least 16 or 17 amino acids, depending on the size of thepeptides of comparison. Accordingly, any of the above-specified levelsof identity may be across the entire length of sequence.

In connection with amino acid sequences, “sequence identity” refers tosequences which have the stated value when assessed using ClustalW(Thompson et al, Nucleic Acids Res. 1994 Nov. 11; 22(22):4673-80) withthe following parameters: Pairwise alignment parameters—Method:accurate, Matrix: PAM, Gap open penalty: 10.00, Gap extension penalty:0.10; Multiple alignment parameters—Matrix: PAM, Gap open penalty:10.00, % identity for delay: 30, Penalize end gaps: on, Gap separationdistance: 0, Negative matrix: no, Gap extension penalty: 0.20,Residue-specific gap penalties: on, Hydrophilic gap penalties: on,Hydrophilic residues: GPSNDQEKR. Sequence identity at a particularresidue is intended to include identical residues which have simply beenderivatised.

A variant peptide may comprise 1, 2, 3, 4, 5 or more, or up to 10 aminoacid substitutions from any of SEQ ID NOS: 1 to 104. Substitutionvariants preferably involve the replacement of one or more amino acidswith the same number of amino acids and making conservative amino acidsubstitutions. For example, an amino acid may be substituted with analternative amino acid having similar properties, for example, anotherbasic amino acid, another acidic amino acid, another neutral amino acid,another charged amino acid, another hydrophilic amino acid, anotherhydrophobic amino acid, another polar amino acid, another aromatic aminoacid or another aliphatic amino acid. Some properties of the 20 mainamino acids which can be used to select suitable substituents are asfollows:

Ala aliphatic, hydrophobic, neutral Cys polar, hydrophobic, neutral Asppolar, hydrophilic, charged (−) Glu polar, hydrophilic, charged (−) Phearomatic, hydrophobic, neutral Gly aliphatic, neutral His aromatic,polar, hydrophilic, charged (+) Ile aliphatic, hydrophobic, neutral Lyspolar, hydrophilic, charged (+) Leu aliphatic, hydrophobic, neutral Methydrophobic, neutral Asn polar, hydrophilic, neutral Pro hydrophobic,neutral Gln polar, hydrophilic, neutral Arg polar, hydrophilic, charged(+) Ser polar, hydrophilic, neutral Thr polar, hydrophilic, neutral Valaliphatic, hydrophobic, neutral Trp aromatic, hydrophobic, neutral Tyraromatic, polar, hydrophobic

Further variants include those in which instead of the naturallyoccurring amino acid the amino acid which appears in the sequence is astructural analog thereof. Amino acids used in the sequences may also bemodified, e.g. labelled, providing the function of the peptide is notsignificantly adversely affected. Where the peptide has a sequence thatvaries from the sequence of any of SEQ ID NOS: 1 to 104 or a fragmentthereof, the substitutions may occur across the full length of thesequence, within the sequence of any of SEQ ID NOS: 1 to 104 or outsidethe sequence of any of SEQ ID NOS: 1 to 104. For example, the variationsdescribed herein, such as additions, deletions, substitutions andmodifications, may occur within the sequence of any of SEQ ID NOS: 1 to104. A variant peptide may comprise or consist essentially of the aminoacid sequence of any of SEQ ID NOS: 1 to 104 in which one, two, three,four or more amino acid substitutions have been made. A variant peptidemay comprise a fragment of the parent protein that is larger than any ofSEQ ID NOS: 1 to 104. In this embodiment, the variations describedherein, such as substitutions and modifications, may occur within and/oroutside the sequence of any of SEQ ID NOS: 1 to 104.

The variant peptides of the invention are 9 to 30 amino acids in lengthinclusive. Preferably, they may be from 9 to 20 or more preferably 13 to17 amino acids in length. The peptides may be the same length as thepeptide sequences in any one of SEQ ID NOS: 1 to 20.

The peptides may be chemically derived from the polypeptide allergen,for example by proteolytic cleavage or can be derived in an intellectualsense from the polypeptide allergen, for example by making use of theamino acid sequence of the polypeptide allergen and synthesisingpeptides based on the sequence. Peptides may be synthesised usingmethods well known in the art.

Where polypeptides comprise residues which are typically difficult topreserve during manufacture, these residues may be replaced. Forexample, glutamate spontaneously forms pyroglutamate in solutionparticularly when present at the N terminus of a peptide. Thus, residuesof the peptides of the invention which correspond to glutamate in thesequence of a native allergen protein sequence may be replaced withpyrogluatmate in the peptides of the invention when such residues arepresent at the N terminus of a peptide.

The term “peptide” includes not only molecules in which amino acidresidues are joined by peptide (—CO—NH—) linkages but also molecules inwhich the peptide bond is reversed. Such retro-inverso peptidomimeticsmay be made using methods known in the art, for example such as thosedescribed in Meziere et al (1997) J. Immunol. 159, 3230-3237. Thisapproach involves making pseudopeptides containing changes involving thebackbone, and not the orientation of side chains. Meziere et al (1997)show that, at least for MHC class II and T helper cell responses, thesepseudopeptides are useful. Retro-inverse peptides, which contain NH—CObonds instead of CO—NH peptide bonds, are much more resistant toproteolysis.

Similarly, the peptide bond may be dispensed with altogether providedthat an appropriate linker moiety which retains the spacing between thecarbon atoms of the amino acid residues is used; it is particularlypreferred if the linker moiety has substantially the same chargedistribution and substantially the same planarity as a peptide bond. Itwill also be appreciated that the peptide may conveniently be blocked atits N- or C-terminus so as to help reduce susceptibility toexoproteolytic digestion. For example, the N-terminal amino group of thepeptides may be protected by reacting with a carboxylic acid and theC-terminal carboxyl group of the peptide may be protected by reactingwith an amine. Other examples of modifications include glycosylation andphosphorylation. Another potential modification is that hydrogens on theside chain amines of R or K may be replaced with methylene groups(—NH₂→—NH(Me) or —N(Me)₂).

Analogues of peptides according to the invention may also includepeptide variants that increase or decrease the peptide's half-life invivo. Examples of analogues capable of increasing the half-life ofpeptides used according to the invention include peptoid analogues ofthe peptides, D-amino acid derivatives of the peptides, andpeptide-peptoid hybrids. A further embodiment of the variantpolypeptides used according to the invention comprises D-amino acidforms of the polypeptide. The preparation of polypeptides using D-aminoacids rather than L-amino acids greatly decreases any unwanted breakdownof such an agent by normal metabolic processes, decreasing the amountsof agent which needs to be administered, along with the frequency of itsadministration.

The peptides provided by the present invention may be derived fromsplice variants of the parent proteins encoded by mRNA generated byalternative splicing of the primary transcripts encoding the parentprotein chains. The peptides may also be derived from amino acidmutants, glycosylation variants and other covalent derivatives of theparent proteins which retain at least an MHC-binding property of theallergens. Exemplary derivatives include molecules wherein the peptidesof the invention are covalently modified by substitution, chemical,enzymatic, or other appropriate means with a moiety other than anaturally occurring amino acid. Further included are naturally occurringvariants of the parent proteins found in different mites. Such a variantmay be encoded by an allelic variant or represent an alternativesplicing variant.

Variants as described above may be prepared during synthesis of thepeptide or by post-production modification, or when the peptide is inrecombinant form using the known techniques of site-directedmutagenesis, random mutagenesis, or enzymatic cleavage and/or ligationof nucleic acids.

In accordance with the invention, the further peptides that thecomposition may comprise are preferably functional variants of any ofSEQ ID NOS: 1 to 104. That is, the peptides are preferably capable ofinducing an immune response. In particular, the peptides are preferablycapable of inducing cytokine production in house dust mite allergicindividuals. Typically, the composition of the invention will thereforecomprise at least one polypeptide or variant thereof which produces acytokine response in greater than 30, 35, 40%, preferably 45% or 50% ofindividuals in a population of house dust mite allergic individuals. Thenumber of individuals in a panel of dust mite allergic individuals maybe any number greater than one, for example at least 20, 30, 40, 50, 80,or at least 100 individuals. Preferably the composition comprises atleast two, three or most preferably four such peptides. Preferably thecytokine response is production of IL13 or IFN-gamma. Cytokineproduction may be measured by any suitable method. Production of acytokine is typically considered to have occurred in response to apeptide if the level of cytokine produced in the presence of the peptideis at least 2, 3, 4 or 5 fold above the background level of saidcytokine that is produced in the absence of a stimulus (i.e. the levelproduced by the same individual in the absence of the peptide or anyother stimulus). Alternatively, production of a cytokine may beconsidered to have occurred if the amount of cytokine produced exceeds arecognised limit, typically 90, 95, or preferably 100 pg/ml, typicallyfrom a sample of approximately 1.25×10⁶ cells in 250 μl.

Suitable methods for measuring cytokine production typically includemeasuring the cytokine release from peripheral blood mononuclear cells(PBMCs) from a taken sample from a subject. The sample is typicallyblood or serum. Cytokine release from PBMCs is measured after incubatingthe cells in the presence of a given peptide. Supernatants from theincubation mixture are then tested for the presence of a cytokine, usingany suitable assay, for example an ELISA, ELISPOT assay or flowcytometric assay. Particularly preferred methods include Multiplex beadarray assays as described in, for example de Jager et al; Clinical andDiagnostic Laboratory Immunology, 2003, Vol 10(1) p. 133-139. Typically,the composition may comprise at least one additional peptide or variantthereof that is not amongst the polypeptides already selected, upto atotal of thirteen different peptides, which produces a cytokine responsein greater than 20%, 25%, preferably 30%, 35% or 40% of individuals in apopulation of house dust mite allergic individuals.

The composition may further comprise one or more additional peptides orvariants thereof that are not amongst the polypeptides already selected,upto a total of thirteen different peptides, which produce a cytokineresponse in greater than 10%, 15%, preferably 20% of individuals in apopulation of house dust mite allergic individuals.

Suitable variants capable of binding to TCRs may be derived empiricallyor selected according to known criteria. Within a single peptide thereare certain residues which contribute to binding within the MHC antigenbinding groove and other residues which interact with hypervariableregions of the T cell receptor (Allen et al (1987) Nature 327: 713-5).

Within the residues contributing to T cell receptor interaction, ahierarchy has been demonstrated which pertains to dependency of T cellactivation upon substitution of a given peptide residue. Using peptideswhich have had one or more T cell receptor contact residues substitutedwith a different amino acid, several groups have demonstrated profoundeffects upon the process of T cell activation. Evavold & Allen (1991)Nature 252: 1308-10) demonstrated the dissociation of T cellproliferation and cytokine production. In this in vitro model, a T cellclone specific for residues 64-76 of haemoglobin (in the context ofI-E^(k)), was challenged with a peptide analogue in which a conservativesubstitution of aspartic acid for glutamic acid had been made. Thissubstitution did not significantly interfere with the capacity of theanalogue to bind to I-E^(k).

Following in vitro challenge of a T cell clone with this analogue, noproliferation was detected although IL-4 secretion was maintained, aswas the capacity of the clone to help B cell responses. In a subsequentstudy the same group demonstrated the separation of T cell-mediatedcytolysis from cytokine production. In this instance, the formerremained unaltered while the latter was impaired. The efficacy ofaltered peptide ligands in vivo was initially demonstrated in a murinemodel of EAE (experimental allergic encephalomyelitis) by McDevitt andcolleagues (Smilek et al (1991) Proc Natl Acad Sci USA 88: 9633-9637).In this model EAE is induced by immunisation with the encephalitogenicpeptide Act-11 of MBP (myelin basic protein). Substitution at positionfour (lysine) with an alanine residue generated a peptide which boundwell to its restricting element (Aα^(u)Aβ^(u)), but which wasnon-immunogenic in the susceptible PL/JxSJLF1 strain and which,furthermore prevented the onset of EAE when administered either beforeor after immunisation with the encephalitogenic peptide. Thus, residuescan be identified in peptides which affect the ability of the peptidesto induce various functions of T-cells.

Advantageously, peptides may be designed to favour T-cell proliferationand induction of desensitisation. Metzler and Wraith have demonstratedimproved tolerogenic capacity of peptides in which substitutionsincreasing peptide-MHC affinity have been made (Metzler & Wraith (1993)Int Immunol˜: 1159-65). That an altered peptide ligand can causelong-term and profound anergy in cloned T cells was demonstrated bySloan-Lancaster et al (1993) Nature 363: 156-9.

The compositions of the invention are capable of inducing a late phaseresponse in an individual that is sensitised to the allergens. The term“late phase response” includes the meaning as set forth in Allergy andAllergic Diseases (1997) A. B. Kay (Ed.), Blackwell Science, pp1113-1130. The late phase response may be any late phase response (LPR).Preferably, the peptides are capable of inducing a late asthmaticresponse (LAR) or a late rhinitic response, or a late phase skinresponse or a late phase ocular response. Whether or not a particularpeptide can give rise to a LPR can be determined using methods wellknown in the art; a particularly preferred method is that described inCromwell O, Durham S R, Shaw R J, Mackay J and Kay A B. Provocationtests and measurements of mediators from mast cells and basophils inasthma and allergic rhinitis. In: Handbook of Experimental Immunology(4) Chapter 127, Editor: Weir D M, Blackwell Scientific Publications,1986.

Thus, preferably, the individual peptides of the invention are able toinduce a LPR in an individual who has been sensitised to the allergens.Whether or not an individual has been sensitised to the allergens may bedetermined by well known procedures such as skin prick testing withsolutions of allergen extracts, induction of cutaneous LPRs, clinicalhistory, allergen challenge and radioallergosorbent test (RAST) formeasurement of allergen specific IgE. Whether or not a particularindividual is expected to benefit from treatment may be determined bythe physician based, for example, on such tests.

Desensitising or tolerising an individual to the allergens meansinhibition or dampening of allergic tissue reactions induced by theallergens in appropriately sensitised individuals. It has been shownthat T cells can be selectively activated, and then renderedunresponsive. Moreover the anergising or elimination of these T-cellsleads to desensitisation of the patient for a particular allergen. Thedesensitisation manifests itself as a reduction in response to anallergen or allergen-derived peptide, or preferably an elimination ofsuch a response, on second and further administrations of the allergenor allergen-derived peptide. The second administration may be made aftera suitable period of time has elapsed to allow desensitisation to occur;this is preferably any period between one day and several weeks. Aninterval of around two weeks is preferred.

Although the compositions of the invention are able to induce a LPR in adust mite allergic individual, it should be appreciated that when acomposition is used to treat a patient it is preferable that asufficiently low concentration of the composition is used such that noobservable LPR will occur but the response will be sufficient topartially desensitise the T cells such that the next (preferably higher)dose may be given, and so on. In this way the dose is built up to givefull desensitisation but often without ever inducing a LPR in thepatient. Although, the composition or peptide is able to do so at ahigher concentration than is administered.

The compositions of the invention preferably are capable of inducing alate phase response in 50% or more of a panel of dust mite allergicindividuals from the population. More preferably, the compositions arecapable of inducing a LPR in 55% or more, 60% or more, 65% or more, 70%or more, 75% or more, 80% or more, 85% or more, or 90% or more ofsensitized individuals in a panel. Whether or not the compositions areable to induce a LPR in a certain percentage of a panel of subjects canbe determined by methods which are well known in the art.

It will be understood that the peptides of the invention comprise a Tcell epitope that consists of a core 9 amino acids which are the minimalessential sequence required for MHC class II binding. However, thepeptides may also comprise additional residues flanking the core 9 aminoacids. The peptides may therefore comprise a region containing a T cellepitope, in which some residues may be modified without affecting thefunction of the epitope. Accordingly, functional variants of thepeptides as defined above include peptides which are altered to improvetheir solubility relative to the native sequence of the peptides.Improved solubility is advantageous for the tolerisation of subjects toallergens from which the peptides of the invention derive, sinceadministration of poorly soluble agents to subjects causes undesirable,non-tolerising inflammatory responses. The solubility of the peptidesmay be improved by altering the residues which flank the regioncontaining a T cell epitope. A peptide of the invention may beengineered to be more soluble such that it comprises:

-   i) N terminal to the residues of the peptide which flank a T cell    epitope: one to six contiguous amino acids corresponding to the two    to six contiguous amino acids immediately N terminal to said    residues in the sequence of the protein from which the peptide    derives; and/or-   ii) C terminal to the residues of the peptide which flank a T cell    epitope: one to six contiguous amino acids corresponding to the one    to six contiguous amino acids immediately C terminal to the said    residues in the sequence of the protein from which the peptide    derives; or-   iii) both N and C terminal to the residues of the peptide which    flank a T cell epitope, at least one amino acid selected from    arginine, lysine, histidine, glutamate and aspartate.

Optionally, the peptides may additionally be engineered to be moresoluble such that:

i) any cysteine residues in the native sequence of the peptide arereplaced with serine or 2-aminobutyric acid; and/or

ii) any residues at the N or C terminus of the native sequence of thepeptide, which are not comprised in a T cell epitope, are deleted;and/or

iii) any two consecutive amino acids comprising the sequence Asp-Gly inthe upto four amino acids at the N or C terminus of the native sequenceof the peptide, which are not comprised in a T cell epitope, aredeleted.

Nucleic Acids and Vectors

The individual peptides that make up the compositions and products ofthe invention may be administered directly, or may be administeredindirectly by expression from an encoding sequence. For example, apolynucleotide may be provided that encodes a peptide of the invention,such as any of the peptides described above. A peptide of the inventionmay thus be produced from or delivered in the form of a polynucleotidewhich encodes, and is capable of expressing, it. Any reference herein tothe use, delivery or administration of a peptide of the invention isintended to include the indirect use, delivery or administration of sucha peptide via expression from a polynucleotide that encodes it.

The terms “nucleic acid molecule” and “polynucleotide” are usedinterchangeably herein and refer to a polymeric form of nucleotides ofany length, either deoxyribonucleotides or ribonucleotides, or analogsthereof. Non-limiting examples of polynucleotides include a gene, a genefragment, messenger RNA (mRNA), cDNA, recombinant polynucleotides,plasmids, vectors, isolated DNA of any sequence, isolated RNA of anysequence, nucleic acid probes, and primers. A polynucleotide of theinvention may be provided in isolated or purified form. A nucleic acidsequence which “encodes” a selected polypeptide is a nucleic acidmolecule which is transcribed (in the case of DNA) and translated (inthe case of mRNA) into a polypeptide in vivo when placed under thecontrol of appropriate regulatory sequences. The boundaries of thecoding sequence are determined by a start codon at the 5′ (amino)terminus and a translation stop codon at the 3′ (carboxy) terminus. Forthe purposes of the invention, such nucleic acid sequences can include,but are not limited to, cDNA from viral, prokaryotic or eukaryotic mRNA,genomic sequences from viral or prokaryotic DNA or RNA, and evensynthetic DNA sequences. A transcription termination sequence may belocated 3′ to the coding sequence.

Polynucleotides of the invention can be synthesised according to methodswell known in the art, as described by way of example in Sambrook et al(19104, Molecular Cloning—a laboratory manual; Cold Spring HarborPress).

The polynucleotide molecules of the present invention may be provided inthe form of an expression cassette which includes control sequencesoperably linked to the inserted sequence, thus allowing for expressionof the peptide of the invention in vivo in a targeted subject. Theseexpression cassettes, in turn, are typically provided within vectors(e.g., plasmids or recombinant viral vectors) which are suitable for useas reagents for nucleic acid immunization. Such an expression cassettemay be administered directly to a host subject. Alternatively, a vectorcomprising a polynucleotide of the invention may be administered to ahost subject. Preferably the polynucleotide is prepared and/oradministered using a genetic vector. A suitable vector may be any vectorwhich is capable of carrying a sufficient amount of genetic information,and allowing expression of a peptide of the invention.

The present invention thus includes expression vectors that comprisesuch polynucleotide sequences. Thus, the present invention provides avector for use in preventing or treating allergy to dust mites bytolerisation comprising four or more polynucleotide sequences whichencode different polypeptides of the invention and optionally one ormore further polynucleotide sequences which encode differentpolypeptides as defined herein. The vector may comprise 4, 5, 6 or 7polynucleotide sequences which encode different polypeptides of theinvention.

Furthermore, it will be appreciated that the compositions and productsof the invention may comprise a mixture of polypeptides andpolynucleotides. Accordingly, the invention provides a composition orproduct as defined herein, wherein in place of any one of thepolypeptide is a polynucleotide capable of expressing said polypeptide.

Expression vectors are routinely constructed in the art of molecularbiology and may for example involve the use of plasmid DNA andappropriate initiators, promoters, enhancers and other elements, such asfor example polyadenylation signals which may be necessary, and whichare positioned in the correct orientation, in order to allow forexpression of a peptide of the invention. Other suitable vectors wouldbe apparent to persons skilled in the art. By way of further example inthis regard we refer to Sambrook et al.

Thus, a polypeptide of the invention may be provided by delivering sucha vector to a cell and allowing transcription from the vector to occur.Preferably, a polynucleotide of the invention or for use in theinvention in a vector is operably linked to a control sequence which iscapable of providing for the expression of the coding sequence by thehost cell, i.e. the vector is an expression vector.

“Operably linked” refers to an arrangement of elements wherein thecomponents so described are configured so as to perform their usualfunction. Thus, a given regulatory sequence, such as a promoter,operably linked to a nucleic acid sequence is capable of effecting theexpression of that sequence when the proper enzymes are present. Thepromoter need not be contiguous with the sequence, so long as itfunctions to direct the expression thereof. Thus, for example,intervening untranslated yet transcribed sequences can be presentbetween the promoter sequence and the nucleic acid sequence and thepromoter sequence can still be considered “operably linked” to thecoding sequence.

A number of expression systems have been described in the art, each ofwhich typically consists of a vector containing a gene or nucleotidesequence of interest operably linked to expression control sequences.These control sequences include transcriptional promoter sequences andtranscriptional start and termination sequences. The vectors of theinvention may be for example, plasmid, virus or phage vectors providedwith an origin of replication, optionally a promoter for the expressionof the said polynucleotide and optionally a regulator of the promoter. A“plasmid” is a vector in the form of an extrachromosomal geneticelement. The vectors may contain one or more selectable marker genes,for example an ampicillin resistance gene in the case of a bacterialplasmid or a resistance gene for a fungal vector. Vectors may be used invitro, for example for the production of DNA or RNA or used to transfector transform a host cell, for example, a mammalian host cell. Thevectors may also be adapted to be used in vivo, for example to allow invivo expression of the polypeptide.

A “promoter” is a nucleotide sequence which initiates and regulatestranscription of a polypeptide-encoding polynucleotide. Promoters caninclude inducible promoters (where expression of a polynucleotidesequence operably linked to the promoter is induced by an analyte,cofactor, regulatory protein, etc.), repressible promoters (whereexpression of a polynucleotide sequence operably linked to the promoteris repressed by an analyte, cofactor, regulatory protein, etc.), andconstitutive promoters. It is intended that the term “promoter” or“control element” includes full-length promoter regions and functional(e.g., controls transcription or translation) segments of these regions.

A polynucleotide, expression cassette or vector according to the presentinvention may additionally comprise a signal peptide sequence. Thesignal peptide sequence is generally inserted in operable linkage withthe promoter such that the signal peptide is expressed and facilitatessecretion of a polypeptide encoded by coding sequence also in operablelinkage with the promoter.

Typically a signal peptide sequence encodes a peptide of 10 to 30 aminoacids for example 15 to 20 amino acids. Often the amino acids arepredominantly hydrophobic. In a typical situation, a signal peptidetargets a growing polypeptide chain bearing the signal peptide to theendoplasmic reticulum of the expressing cell. The signal peptide iscleaved off in the endoplasmic reticulum, allowing for secretion of thepolypeptide via the Golgi apparatus. Thus, a peptide of the inventionmay be provided to an individual by expression from cells within theindividual, and secretion from those cells.

Alternatively, polynucleotides of the invention may be expressed in asuitable manner to allow presentation of a peptide of the invention byan MHC class II molecule at the surface of an antigen presenting cell.For example, a polynucleotide, expression cassette or vector of theinvention may be targeted to antigen presenting cells, or the expressionof encoded peptide may be preferentially stimulated or induced in suchcells.

Polynucleotides of interest may be used in vitro, ex vivo or in vivo inthe production of a peptide of the invention. Such polynucleotides maybe administered or used in the prevention or treatment of allergy bytolerisation.

Methods for gene delivery are known in the art. See, e.g., U.S. Pat.Nos. 5,399,346, 5,580,859 and 5,5104,466. The nucleic acid molecule canbe introduced directly into the recipient subject, such as by standardintramuscular or intradermal injection; transdermal particle delivery;inhalation; topically, or by oral, intranasal or mucosal modes ofadministration. The molecule alternatively can be introduced ex vivointo cells that have been removed from a subject. For example, apolynucleotide, expression cassette or vector of the invention may beintroduced into APCs of an individual ex vivo. Cells containing thenucleic acid molecule of interest are re-introduced into the subjectsuch that an immune response can be mounted against the peptide encodedby the nucleic acid molecule. The nucleic acid molecules used in suchimmunization are generally referred to herein as “nucleic acidvaccines.”

The polypeptides, polynucleotides, vectors or cells of the invention maybe present in a substantially isolated form. They may be mixed withcarriers or diluents which will not interfere with their intended useand still be regarded as substantially isolated. They may also be in asubstantially purified form, in which case they will generally compriseat least 90%, e.g. at least 95%, 98% or 99%, of the proteins,polynucleotides, cells or dry mass of the preparation.

Antigen Presenting Cells (APCs)

The invention encompasses the use in vitro of a method of producing apopulation of APCs that present the peptides of the invention on theirsurface, that may be subsequently used in therapy. Such a method may becarried out ex vivo on a sample of cells that have been obtained from apatient. The APCs produced in this way therefore form a pharmaceuticalagent that can be used in the treatment or prevention of dust miteallergy by tolerisation. The cells should be accepted by the immunesystem of the individual because they derive from that individual.Delivery of cells that have been produced in this way to the individualfrom whom they were originally obtained, thus forms a therapeuticembodiment of the invention.

Formulations and Compositions

The peptides, polynucleotides, vectors and cells of the invention may beprovided to an individual either singly or in combination. Each moleculeor cell of the invention may be provided to an individual in anisolated, substantially isolated, purified or substantially purifiedform. For example, a peptide of the invention may be provided to anindividual substantially free from the other peptides.

Whilst it may be possible for the peptides, polynucleotides orcompositions according to the invention to be presented in raw form, itis preferable to present them as a pharmaceutical formulation. Thus,according to a further aspect of the invention, the present inventionprovides a pharmaceutical formulation for use in preventing or treatingallergy to dust mites by tolerisation comprising a composition, vectoror product according to the invention together with one or morepharmaceutically acceptable carriers or diluents and optionally one ormore other therapeutic ingredients. The carrier (s) must be ‘acceptable’in the sense of being compatible with the other ingredients of theformulation and not deleterious to the recipient thereof. Typically,carriers for injection, and the final formulation, are sterile andpyrogen free.

Formulation of a composition comprising the peptide, polynucleotides orcells of the invention can be carried out using standard pharmaceuticalformulation chemistries and methodologies all of which are readilyavailable to the reasonably skilled artisan.

For example, compositions containing one or more molecules or cells ofthe invention can be combined with one or more pharmaceuticallyacceptable excipients or vehicles. Auxiliary substances, such as wettingor emulsifying agents, pH buffering substances and the like, may bepresent in the excipient or vehicle. These excipients, vehicles andauxiliary substances are generally pharmaceutical agents that do notinduce an immune response in the individual receiving the composition,and which may be administered without undue toxicity. Pharmaceuticallyacceptable excipients include, but are not limited to, liquids such aswater, saline, polyethyleneglycol, hyaluronic acid and ethanol.Pharmaceutically acceptable salts can also be included therein, forexample, mineral acid salts such as hydrochlorides, hydrobromides,phosphates, sulfates, and the like; and the salts of organic acids suchas acetates, propionates, malonates, benzoates, and the like. A thoroughdiscussion of pharmaceutically acceptable excipients, vehicles andauxiliary substances is available in Remington's Pharmaceutical Sciences(Mack Pub. Co., N.J. 1991).

Such compositions may be prepared, packaged, or sold in a form suitablefor bolus administration or for continuous administration. Injectablecompositions may be prepared, packaged, or sold in unit dosage form,such as in ampoules or in multi-dose containers containing apreservative. Compositions include, but are not limited to, suspensions,solutions, emulsions in oily or aqueous vehicles, pastes, andimplantable sustained-release or biodegradable formulations. Suchcompositions may further comprise one or more additional ingredientsincluding, but not limited to, suspending, stabilizing, or dispersingagents. In one embodiment of a composition for parenteraladministration, the active ingredient is provided in dry (for e.g., apowder or granules) form for reconstitution with a suitable vehicle (e.g., sterile pyrogen-free water) prior to parenteral administration ofthe reconstituted composition. The pharmaceutical compositions may beprepared, packaged, or sold in the form of a sterile injectable aqueousor oily suspension or solution. This suspension or solution may beformulated according to the known art, and may comprise, in addition tothe active ingredient, additional ingredients such as the dispersingagents, wetting agents, or suspending agents described herein. Suchsterile injectable formulations may be prepared using a non-toxicparenterally-acceptable diluent or solvent, such as water or 1,3-butanediol, for example. Other acceptable diluents and solvents include, butare not limited to, Ringer's solution, isotonic sodium chloridesolution, and fixed oils such as synthetic mono- or di-glycerides. Otherparentally-administrable compositions which are useful include thosewhich comprise the active ingredient in microcrystalline form, in aliposomal preparation, or as a component of a biodegradable polymersystems. Compositions for sustained release or implantation may comprisepharmaceutically acceptable polymeric or hydrophobic materials such asan emulsion, an ion exchange resin, a sparingly soluble polymer, or asparingly soluble salt.

Alternatively, the peptides or polynucleotides of the present inventionmay be encapsulated, adsorbed to, or associated with, particulatecarriers. Suitable particulate carriers include those derived frompolymethyl methacrylate polymers, as well as PLG microparticles derivedfrom poly(lactides) and poly(lactide-co-glycolides). See, e.g., Jefferyet al. (1993) Pharm. Res. 10:362-368. Other particulate systems andpolymers can also be used, for example, polymers such as polylysine,polyarginine, polyornithine, spermine, spermidine, as well as conjugatesof these molecules.

The formulation of any of the peptides, polynucleotides or cellsmentioned herein will depend upon factors such as the nature of thesubstance and the method of delivery. Any such substance may beadministered in a variety of dosage forms. It may be administered orally(e.g. as tablets, troches, lozenges, aqueous or oily suspensions,dispersible powders or granules), parenterally, epicutaneously,subcutaneously, by inhalation, intravenously, intramuscularly,intrasternally, transdermally, intradermally, sublingually,instranasally, buccally or by infusion techniques. The substance mayalso be administered as suppositories. A physician will be able todetermine the required route of administration for each particularindividual.

The compositions of formulations of the invention will comprise asuitable concentration of each peptide/polynucleotide/cell to beeffective without causing adverse reaction. Typically, the concentrationof each peptide in the composition will be in the range of 0.03 to 200nmol/ml. More preferably in the range of 0.3 to 200 nmol/ml, 3 to 180nmol/ml, 10 to 150 nmol/ml or 30 to 120 nmol/ml. The composition orformulations should have a purity of greater than 95% or 98% or a purityof at least 99%.

In one embodiment, therefore, the peptides, polynucleotides, cells orcompositions of the invention are used for therapy in combination withone or more other therapeutic agents. The agents may be administeredseparately, simultaneously or sequentially. They may be administered inthe same or different compositions. Accordingly, in a method of theinvention, the subject may also be treated with a further therapeuticagent.

A composition may therefore be formulated which comprises a moleculeand/or cell of the invention and also one or more other therapeuticmolecules. A composition of the invention may alternatively be usedsimultaneously, sequentially or separately with one or more othertherapeutic compositions as part of a combined treatment.

Therapeutic Methods and Individual to be Treated

The present invention relates to peptides, polynucleotides, vectors andcells that are capable of desensitising or tolerising human individualsto the allergens described above and are therefore useful in theprevention or treatment of dust mite allergy. The invention providescompositions, products, vectors and formulations for use in preventingor treating allergy to dust mites by tolerisation. The invention alsoprovides a method of tolerising or desensitizing a dust mite allergicindividual comprising administering, either singly or in combination thepolypeptides/polynucleotides/cells of the invention as described above.

The individual to be treated or provided with the composition orformulation of the invention is preferably human. It will be appreciatedthat the individual to be treated may be known to be sensitised to theallergens, at risk of being sensitised or suspected of being sensitised.The individual can be tested for sensitisation using techniques wellknown in the art and as described herein. Alternatively, the individualmay have a family history of allergy to dust mites. It may not benecessary to test an individual for sensitisation to dust mites becausethe individual may display symptoms of allergy when exposed to dustmites. By exposure is meant proximity to, for example, an item ofclothing, a mattress, pillow, pillow case, sheet, blanket or otherbedding material which has not been washed at greater than 50° C. formore than approximately one week, or a carpet, curtain or upholstereditem of furniture which has not been vacuum cleaned for more thanapproximately one week. By proximity is meant 10 meters or less, 5meters or less, 2 meters or less, 1 meter or less, or 0 meters from theitems described above. Symptoms of allergy can include itchy eyes, runnynose, breathing difficulties, red itchy skin or rash.

The individual to be treated may be of any age. However, preferably, theindividual may be in the age group of 1 to 90, 5 to 60, 10 to 40, ormore preferably 18 to 35.

Preferably, the individual to be treated is from a population that hasMHC allele frequencies within the range of frequencies that arerepresentative of the Caucasian population. Reference population allelefrequencies for 11 common DRB1 allele families are shown in Table 1(Data from HLA Facts Book, Parham and Barber).

TABLE 1 DRB1 1 3 4 7 8 11 12 13 14 15 16 % 6.4 14.7 15.7 8.8 3.4 8.3 3.914.7 2.9 17.6 2.5 Reference population % 9.4 11.1 12.8 13.2 3.7 13.4 2.310.2 3.2 10.7 3.6Reference frequencies were obtained by analysis of multiple studiesreporting frequencies and the figures shown are mean values. Preferablytherefore, the individual to be treated is from a population that hasequivalent MHC allele frequencies as the reference population for thealleles referred to Table 1 (such as for at least 1, 2, 3, 4, 5 or allof the alleles), for example within the ranges of those figures plus orminus 1, 2, 3, 5, 10, 15 or 20%.

Preferably the individual is from a population where the allelefrequencies of the following DRB1 alleles is:

4—at least 9%

7—at least 10%

11—at least 8%.

The individual may have had allergy to dust mites for at least 2 weeks,1 month, 6 months, 1 year or 5 years. The individual may suffer from arash, nasal congestion, nasal discharge and/or coughing caused by theallergy. The individual may or may not have been administered with othercompositions/compounds which treat dust mite allergy. The individual maylive in a geographical region which experiences a daily average relativehumidity greater than 50%, preferably 55%, 60%, 65%, 70%, 75%, 80% or90%. The individual may live in a geographical region known to supportdust mite populations, for example the eastern half of the United States(and major western coastal cities of the United States), populous areasof Canada, western Europe, Japan, Korea, and coastal areas of SouthAmerica, Australia and South Africa.

Combination Immunotherapy

Since many individuals are allergic, or may require desensitizing toseveral polypeptide antigens, the current invention also provides meansof desensitizing individuals that are allergic to multiple antigens.“Tolerance” induced in an individual to a first polypeptide antigen orallergen can create in the individual a “tolergeneic environment”wherein inappropriate immune responses to other antigens can bedownregulated in order to provide tolerance to other antigens.

This finding means that individuals allergic to multiple allergens canbe treated in a greatly reduced time period, and that individualsseriously allergic to some allergens (e.g., peanuts) but more mildlyallergic to other allergens (e.g., cat dander) can benefit from atherapy wherein tolerance to the milder allergen is established and thenthis tolergeneic environment is used to provide tolerance to the other,more extreme allergen. In addition, individuals suffering from anautoimmune disorder who are additionally sensitised (or otherwiseimmune) to an unrelated antigen or allergen can benefit from a treatmentregime wherein tolerance to the unrelated antigen or allergen is firstestablished and then this tolergeneic environment is used to providetolerance to the autoantigen associated with the autoimmune disorder.

A method is therefore provided for desensitising a dust mite allergicindividual to dust mite allergen as described above and one or morefurther different polypeptide antigens. The method entails, in a firststep, administering to the individual a composition/product/formulation(primary composition) according to the invention as described herein andwherein the administration is carried out in a manner sufficient togenerate a hyporesponsive state against dust mite allergen. Once ahyporesponsive state has been established toward dust mite allergen, orat least a shift toward desensitisation has occurred, the method entailsadministration of a secondary composition comprising a second, differentpolypeptide antigen to which the individual is to be sensitised.Administration of the secondary composition is carried out in such a wayas to take advantage of the tolergeneic environment established by useof the primary composition, where it is now possible to establishtolerance to the second, different polypeptide antigen. The secondarycomposition is coadministered with either the first primary compositionor a larger fragment of Feld1. By “coadministered” it is meant eitherthe simultaneous or concurrent administration, e.g., when the two arepresent in the same composition or administered in separate compositionsat nearly the same time but at different sites, as well as the deliveryof polypeptide antigens in separate compositions at different times. Forexample, the secondary composition may be delivered prior to orsubsequent to delivery of the first composition at the same or adifferent site. The timing between deliveries can range from aboutseveral seconds apart to about several minutes apart, several hoursapart, or even several days apart. Furthermore, different deliverymethods can be employed.

The second polypeptide antigen is preferably an allergen different tothe dust mite allergen. Suitable allergens for use in the methods of theinvention can of course be obtained and/or produced using known methods.Classes of suitable allergens include, but are not limited to, otherdust mite allergens, pollens, animal dander (especially cat dander),grasses, molds, dusts, antibiotics, stinging insect venoms, and avariety of environmental (including chemicals and metals), drug and foodallergens. Common tree allergens include pollens from cottonwood,popular, ash, birch, maple, oak, elm, hickory, and pecan trees; commonplant allergens include those from mugwort, ragweed, English plantain,sorrel-dock and pigweed; plant contact allergens include those frompoison oak, poison ivy and nettles; common grass allergens include ryegrass, Timothy, Johnson, Bermuda, fescue and bluegrass allergens; commonallergens can also be obtained from molds or fungi such as Alternaria,Fusarium, Hormodendrum, Aspergillus, Micropolyspora, Mucor andthermophilic actinomycetes; epidermal allergens can be obtained fromhouse or organic dusts (typically fungal in origin), or from animalsources such as feathers, and dog dander; common food allergens includemilk and cheese (diary), egg, wheat, nut (e.g., peanut), seafood (e.g.,shellfish), pea, bean and gluten allergens; common environmentalallergens include metals (nickel and gold), chemicals (formaldehyde,trinitrophenol and turpentine), Latex, rubber, fiber (cotton or wool),burlap, hair dye, cosmetic, detergent and perfume allergens; common drugallergens include local anesthetic and salicylate allergens; antibioticallergens include penicillin, tetracycline and sulfonamide allergens;and common insect allergens include bee, wasp and ant venom, andcockroach calyx allergens. Particularly well characterized allergensinclude, but are not limited to, the major cat allergen Fel dl, beevenom phospholipase A2 (PLA) (Akdis et al. (1996) J. Clin. Invest.98:1676-1683), birch pollen allergen Bet v 1 (Bauer et al. (1997) Clin.Exp. Immunol. 107:536-541), and the multi-epitopic recombinant grassallergen rKBG8.3 (Cao et al. (1997) Immunology 90:46-51). These andother suitable allergens are commercially available and/or can bereadily prepared as extracts following known techniques.

Preferably, the second polypeptide allergen is selected from the list ofallergen sequences and database accession numbers (NCBI Entrez accessionnumbers) below. NCBI is the National Center for Biotechnologyinformation and is a division of the US National Institutes of Health.The NCBI web site, from which access to the database may be sought, iswww.ncbi.nlm.nih.gov/. Allergen sequences and database accession numbers(NCBI Entrez accession numbers):

House Dust Mite

Dermatophagoides pteronyssinus

Der p 1 (SEQ ID NO: 143)MKIVLAIASLLALSAVYARPSSIKTFEEYKKAFNKSYATFEDEEAARKNFLESVKYVQSNGGAINHLSDLSLDEFKNRFLMSAEAFEHLKTQFDLNAETNACSINGNAPAEIDLRQMRTVTPIRMQGGCGSCWAFSGVAATESAYLAYRNQSLDLAEQELVDCASQHGCHGDTIPRGIEYIQHNGVVQESYYRYVAREQSCRRPNAQRFGISNYCQIYPPNVNKIREALAQTHSAIAVIIGIKDLDAFRHYDGRTIIQRDNGYQPNYHAVNIVGYSNAQGVDYWIVRNSWDTNWGDNGYGYFAANIDLMMIEEYPYVVIL

Der p 2 (SEQ ID NO: 144)MMYKILCLSLLVAAVARDQVDVKDCANHEIKKVLVPGCHGSEPCIIHRGKPFQLEAVFEANQNTKTAKIEIKASIDGLEVDVPGIDPNACHYMKCPLVKGQQYDIKYTWNVPKIAPKSENVVVTVKVMGDDGVLACAIATHAKIRD

Der p 3 (SEQ ID NO: 145)MITYNILIVLLLAINTLANPILPASPNATIVGGEKALAGECPYQISLQSSSHFCGGTILDEYWILTAAHCVAGQTASKLSIRYNSLKHSLGGEKISVAKIFAHEKYDSYQIDNDIALIKLKSPMKLNQKNAKAVGLPAKGSDVKVGDQVRVSGWGYLEEGSYSLPSELRRVDIAVVSRKECNELYSKANAEVTDNMICGGDVANGGKDSCQGDSGGPVVDVKNNQVVGIVSWGYGCARKGYPGVYTRVGN FIDWIESKRSQ

Der p 4 (SEQ ID NO: 146) KYXTNPHFIGXRSVITXLME

Der p 5 (SEQ ID NO: 147)MKFIIAFFVATLAVMTVSGEDKKHDYQNEFDFLLMERIHEQIKKGELALFYLQEQINHFEEKPTKEMKDKIVAEMDTIIAMIDGVRGVLDRLMQRKDLDIFEQYNLEMAKKSGDILERDLKKEEARVKKIEV

Der p 6 (SEQ ID NO: 148) AIGXQPAAEAEAPFQISLMK

Der p 7 (SEQ ID NO: 149)MMKLLLIAAAAFVAVSADPIHYDKITEEINKAVDEAVAAIEKSETFDPMKVPDHSDKFERHIGIIDLKGELDMRNIQVRGLKQMKRVGDANVKSEDGVVKAHLLVGVHDDVVSMEYDLAYKLGDLHPNTHVISDIQDFVVELSLEVSEEGNMTLTSFEVRQFANVVNHIGGLSILDPIFAVLSDVLTAIFQDTVRAEMTK VLAPAFKKELERNNQ

Der p9 (SEQ ID NO: 150) IVGGSNASPGDAVYQIALDermatophagoides farinae

Der f 1 (SEQ ID NO: 151)MKFVLAIASLLVLTVYARPASIKTFEFKKAFNKNYATVEEEEVARKNFLESLKYVEANKGAINHLSDLSLDEFKNRYLMSAEAFEQLKTQFDLNAETSACRINSVNVPSELDLRSLRTVTPIRMQGGCGSCWAFSGVAATESAYLAYRNTSLDLSEQELVDCASQHGCHGDTIPRGIEYIQQNGVVEERSYPYVAREQRCRRPNSQHYGISNYCQIYPPDVKQIREALTQTHTAIAVIIGIKDLRAFQHYDGRTIIQHDNGYQPNYHAVNIVGYGSTQGDDYWIVRNSWDTTWGDSGYGY FQAGNNLMMIEQYPYVVIM

Der f 2 (SEQ ID NO: 152)MISKILCLSLLVAAVVADQVDVKDCANNEIKKVMVDGCHGSDPCIIHRGKPFTLEALFDANQNTKTAKIEIKASLDGLEIDVPGIDTNACHFMKCPLVKGQQYDIKYTWNVPKIAPKSENVVVTVKLIGDNGVLACAIATHGKIRD

Der f 3 (SEQ ID NO: 153)MMILTIVVLLAANILATPILPSSPNATIVGGVKAQAGDCPYQISLQSSSHFCGGSILDEYWILTAAHCVNGQSAKKLSIRYNTLKHASGGEKIQVAEIYQHENYDSMTIDNDVALIKLKTPMTLDQTNAKPVPLPAQGSDVKVGDKIRVSGWGYLQEGSYSLPSELQRVDIDVVSREQCDQLYSKAGADVSENMICGGDVANGGVDSCQGDSGGPVVDVATKQIVGIVSWGYGCARKGYPGVYTRVGNFV DWIESKRSQ

Der f 4 (SEQ ID NO: 154) AVGGQDADLAEAPFQISLLK

Der f 7 (SEQ ID NO: 155)MMKFLLIAAVAFVAVSADPIHYDKITEEINKAIDDAIAAIEQSETIDPMKVPDHADKFERHVGIVDFKGELAMRNIEARGLKQMKRQGDANVKGEEGIVKAHLLIGVHDDIVSMEYDLAYKLGDLHPTTHVISDIQDFVVALSLEISDEGNITMTSFEVRQFANVVNHIGGLSILDPIFGVLSDVLTAIFQDTVRKEMTK VLAPAFKRELEKNAdditional Mite Allergen Sequences (NCBI Entrez Accession):1170095; 1359436; 2440053; 666007; 487661; 1545803; 84702; 84699;625532; 404370; 1091577; 1460058; 7413; 9072; 387592.CatFelis Sequences (NCBI Entrez Accession):539716; 539715; 423193; 423192; 423191; 423190; 1364213; 1364212;395407; 163827; 163823; 163825; 1169665; 232086; 1169666.LatexHevea Sequences:

Hev b 1 (SEQ ID NO: 156)MAEDEDNQQGQGEGLKYLGFVQDAATYAVTTFSNVYLFAKDKSGPLQPGVDIIEGPVKNVAVPLYNRFSYIPNGALKFVDSTVVASVTIIDRSLPPIVKDASIQVVSAIRAAPEAARSLASSLPGQTKILAKVFYGEN

Hev b 3 (SEQ ID NO: 157)MAEEVEEERLKYLDFVRAAGVYAVDSFSTLYLYAKDISGPLKPGVDTIENVVKTVVTPVYYIPLEAVKFVDKTVDVSVTSLDGVVPPVIKQVSAQTYSVAQDAPRIVLDVASSVFNTGVQEGAKALYANLEPKAEQYAVITWRALNKLPLVPQVANVVVPTAVYFSEKYNDVVRGTTEQGYRVSSYLPLLPTEKITKVFG DEASAdditional Hevea Sequences (NCBI Entrez Accession):3319923; 3319921; 3087805; 1493836; 1480457; 1223884; 3452147; 3451147;1916805; 232267; 123335; 2501578; 3319662; 3288200; 1942537; 2392631;2392630; 1421554; 1311006; 494093; 3183706; 3172534; 283243; 1170248;1708278; 1706547; 464775; 2661042; 231586; 123337; 116359; 123062;2213877; 542013; 2144920; 1070656; 2129914; 2129913; 2129912; 100135;82026; 1076559; 82028; 82027; 282933; 280399; 100138; 1086972; 108697;1086976; 1086978; 1086978; 1086976; 1086974; 1086972; 913758; 913757;913756; 234388; 1092500; 228691; 1177405; 18839; 18837; 18835; 18833;18831; 1209317; 1184668; 168217; 168215; 168213; 168211; 168209; 348137.Rye GrassLolium Sequences:

126385 Lol p 1 (SEQ ID NO: 158)MASSSSVLLVVALFAVFLGSAHGIAKVPPGPNITAEYGDKWLDAKSTWYGKPTGAGPKDNGGACGYKNVDKAPFNGMTGCGNTPIFKDGRGCGSCFEIKCTKPESCSGEAVTVTITDDNEEPIAPYHFDLSGHAFGSMAKKGEEQNVRSAGELELQFRRVKCKYPDDTKPTFHVEKASNPNYLAILVKYVDGDGDVVAVDIKEKGKDKWIELKESWGAVWRIDTPDKLTGPFTVRYTTEGGTKSEFEDVI PEGWKADTSYSAK

126386 Lol p 2a (SEQ ID NO: 159)AAPVEFTVEKGSDEKNLALSIKYNKEGDSMAEVELKEHGSNEWLALKKNGDGVWEIKSDKPLKGPFNFRFVSEKGMRNVFDDVVPADFKVGTTYKPE

126387 Lol p 3 (SEQ ID NO: 160)TKVDLTVEKGSDAKTLVLNIKYTRPGDTLAEVELRQHGSEEWEPMTKKGNLWEVKSAKPLTGPMNFRFLSKGGMKNVFDEVIPTAFTVGKTYTPEYN

2498581 Lol p 5a (SEQ ID NO: 161)MAVQKYTVALFLRRGPRGGPGRSYAADAGYTPAAAATPATPAATPAGGWREGDDRRAEAAGGRQRLASRQPWPPLPTPLRRTSSRSSRPPSPSPPRASSPTSAAKAPGLIPKLDTAYDVAYKAAEAHPRGQVRRLRHCPHRSLRVIAGALEVHAVKPATEEVLAAKIPTGELQIVDKIDAAFKIAATAANAAPTNDKFTVFESAFNKALNECTGGAMRPTSSSPPSRPRSSRPTPPPSPAAPEVKYAVFEAALTKAITAMTQAQKAGKPAAAAATAAATVATAAATAAAVLPPPLLVVQS LISLLIYY

2498582 Lol p 5b (SEQ ID NO: 162)MAVQKHTVALFLAVALVAGPAASYAADAGYAPATPATPAAPATAATPATPATPATPAAVPSGKATTEEQKLIEKINAGFKAAVAAAAVVPPADKYKTFVETFGTATNKAFVEGLASGYADQSKNQLTSKLDAALKLAYEAAQGATPEAKYDAYVATLTEALRVIAGTLEVHAVKPAAEEVKVGAIPAAEVQLIDKVDAAYRTAATAANAAPANDKFTVFENTFNNAIKVSLGAAYDSYKFIPTLVAAVKQAYAAKQATAPEVKYTVSETALKKAVTAMSEAEKEATPAAAATATPTPAAATATATPAAAYATATPAAATATATPAAATATPAAAGGYKV

455288 Lol p isoform 9 (SEQ ID NO: 162)MAVQKHTVALFLAVALVAGPAASYAADAGYAPATPATPAAPATAATPATPATPATPAAVPSGKATTEEQKLIEKINAGFKAAVAAAAVVPPADKYKTFVETFGTATNKAFVEGLASGYADQSKNQLTSKLDAALKLAYEAAQGATPEAKYDAYVATLTEALRVIAGTLEVHAVKPAAEEVKVGAIPAAEVQLIDKVDAAYRTAATAANAAPANDKFTVFENTFNNAIKVSLGAAYDSYKFIPTLVAAVKQAYAAKQATAPEVKYTVSETALKKAVTAMSEAEKEATPAAAATATPTPAAATATATPAAAYATATPAAATATATPAAATATPAAAGGYKV

1582249 Lol p 11 (SEQ ID NO: 164)DKGPGFVVTGRVYCDPCRAGFETNVSHNVEGATVAVDCRPFDGGESKLKAEATTDKDGWYKIEIDQDHQEEICEVVLAKSPDKSCSEIEEFRDRARVPLTSNXGIKQQGIRYANPIAFFRKEPLKECGGILQAYAdditional Lolium Sequences (NCBI Entrez Accession):135480; 417103; 687261; 687259; 1771355; 2388662; 631955; 542131;542130; 542129; 100636; 626029; 542132; 320616; 320615; 320614; 100638;100634; 82450; 626028; 100639; 283345; 542133; 1771353; 1763163;1040877; 1040875; 250525; 551047; 515377; 510911; 939932; 439950; 2718;168316; 168314; 485371; 2388664; 2832717; 2828273; 548867.Olive TreeOlive Sequences

416610 Ole e 1 (SEQ ID NO: 165)EDIPQPPVSOFHIQGQVYCDTCRAGFITELSEFIPGASLRLQCKDKENGDVTFTEVGYTRAEGLYSMLVERDHKNEFCEITLISSGRKDCNEIPTEGWAKPSLKFKLNTVNGTTRTVNPLGFFKKEALPKCAQVYNKLGMYPPNMParietariaParietaria Sequences:

2497750 Par j P2 (SEQ ID NO: 166)MRTVSMAALVVIAAALAWTSSAEPAPAPAPGEEACGKVVQDIMPCLHFVKGEEKEPSKECCSGTKKLSEEVKTTEQKREACKCIVRATKGISGIKNELVAEVPKKCDIKTTLPPITADFDCSKIQSTIFRGYY

1352506 Par j P5 (SEQ ID NO: 167)MVRALMPCLPFVQGKEKEPSKGCCSGAKRLDGETKTGPQRVHACECIQTAMKTYSDIDGKLVSEVPKHCGIVDSKLPPIDVNMDCKTVGVVPRQPQLPVSLRHGPVTGPSDPAHKARLERPOIRVPPPAPEKA

1532056 Par j P8 (SEQ ID NO: 168)MRTVSMAALVVIAAALAWTSSAELASAPAPGEGPCGKVVHHIMPCLKFVKGEEKEPSKSCCSGTKKLSEEVKTTEQKREACKCIVAATKGISGIKNELVAEVPKKCGITTTLPP1TADFDCSKIESTIFRGYY

1532058 Par j P9 (SEQ ID NO: 169)MRTVSAPSAVALVVIVAAGLAWTSLASVAPPAPAPGSEETCGTVVRALMPCLPFVQGKEKEPSKGCCSGAKRLDGETKTGLQRVHACECIQTAMKTYSDIDGKLVSEVPKHCGIVDSKLPPIDVNMDCKTLGVVPRQPQLPVSLRHGPVTGPSDPAHKARLERPQIRVPPPAPEKA

2497749 Par j P9 (SEQ ID NO: 170)MRTVSARSSVALVVIVAAVLVWTSSASVAPAPAPGSEETCGTVVGALMPCLPFVQGKEKEPSKGCCSGAKRLDGETKTGPQRVHACECIQTAMKTYSDIDGKLVSEVPKHCGIVDSKLPPTDVNMDCKTLGVLHYKGN

1086003 Par j 1 (SEQ ID NO: 171)MVRALMPCLPFVQGKEKEPSKGCCSGAKRLDGETKTGPQRVHACECIQTAMKTYSDIDGKLVSEVPKHCGIVDSKLPPIDVNMDCKTVGVVPRQPQLPVSLRHGPVTGPSRSRPPTKHGWRDPRLEFRPPHRKKPNPAFSTLGAdditional Parietaria Sequences (NCBI Entrez Accession):543659; 1836011; 1836010; 1311513; 1311512; 1311511; 1311510; 1311509;240971.Timothy GrassPhleum Sequences:

Phl p 1 (SEQ ID NO: 172)MASSSSVLLVVVLFAVFLGSAYGIPKVPPGPNITATYGDKWLDAKSTWYGKPTGAGPKDNGGACGYKDVDKPPFSGMTGCGNTPIFKSGRGCGSCFEIKCTKPEACSGEPVVVHITDDNEEPIAPYHFDLSGHAFGAMAKKGDEQKLRSAGELELQFRRVKCKYPEGTKVTFHVEKGSNPNYLALLVKYVNGDGDVVAVDIKEKGKDKWIELKESWGAIWRIDTPDKLTGPFTVRYTTEGGTKTEAEDVI PEGWKADTSYESK

Phl p 1 (SEQ ID NO: 173)MASSSSVLLVVALFAVFLGSAHGIPKVPPGPNITATYGDKWLDAKSTWYGKPTAAGPKDNGGACGYKDVDKPPFSGMTGCGNTPIFKSGRGCGSCFEIKCTKPEACSGEPVVVHITDDNEEPIAAYHFDLSGIAFGSMAKKGDEQKLRSAGEVEIQFRRVKCKYPEGTKVTFHVEKGSNPNYLALLVKFSGDGDVVAVDIKEKGKDKWIALKESWGAIWRIDTPEVLKGPFTVRYTTEGGTKARAKDVIP EGWKADTAYESK

Phl p 2 (SEQ ID NO: 174)MSMASSSSSSLLAMAVLAALFAGAWCVPKVTFTVEKGSNEKHLAVLVKYEGDTMAEVELREHGSDEWVAMTKGEGGVWTFDSEEPLQGPFNFRFLTEKGMMKNVFDDVVPEKYTIGATYAPEE

Phl p 5 (SEQ ID NO: 175)ADLGYGGPATPAAPAEAAPAGKATTEEQKLIEKINDGFKAALAAAAGVPPADKYKTFVATFGAASNKAFAEGLSAEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDSAFKVAATAANAAPANDKFTVFEAAFNNAIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAFTAMSEAQKAAKPATEATATATAAVGAATGAATAATGGYKV

Phl p 5 (SEQ ID NO: 176)ADLGYGGPATPAAPAEAAPAGKATTEEQKLIEKINDGFKAALAAAAGVPPADKYKTFVATFGAASNKAFAEGLSAEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDSAFKVAATAANAAPANDKFTVFEAAFNNAIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAITAMSEAQKAAKPATEATATATAAVGAATGAATAATGGYKV

Phl p 5b (SEQ ID NO: 177)AAAAVPRRGPRGGPGRSYTADAGYAPATPAAAGAAAGKATTEEQKLIEDINVGFKAAVAAAASVPAADKFKTFEAAFTSSSKAAAAKAPGLVPKLDAAYSVAYKAAVGATPEAKFDSFVASLTEALRVIAGALEVHAVKPVTEEPGMAKIPAGELQIIDKIDAAFKVAATAAATAPADDKFTVFEAAFNKAIKESTGGAYDTYKCIPSLEAAVKQAYAATVAAAPQVKYAVFEAALTKAITAMSEVQKVSQPATGAATVAAGAATTAAGAASGAATVAAGGYKV

Phl p 5a (SEQ ID NO: 178)ADLGYGPATPAAPAAGYTPATPAAPAGADAAGKATTEEQKLIEKINAGFKAALAGAGVQPADKYRTFVATFGPASNKAFAEGLSGEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDAAFKVAATAANAAPANDKFTVFEAAFNDEIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAITAMSEAQKAAKPAAAATATATAAVGAATGAATAATGGYKV

Phl p 5 (SEQ ID NO: 179)MAVQKYTVALFLAVALVAGPAASYAADAGYAPATPAAAGAEAGKATTEEQKLIEDINVGFKAAVAAAASVPAADKFKTFEAAFTSSSKAATAKAPGLVPKLDAAYSVSYKAAVGATPEAKFDSFVASLTEALRVIAGALEVHAVKPVTEEPGMAKIPAGELQIIDKIDAAFKVAATAAATAPADTVFEAAFNKAIKESTGGAYDTYKCIPSLEAAVKQAYAATVAAAPQVKYAVFEAALTKAITAMSEVQKVSQPATGAATVAAGAATTAAGAASGAATVAAGGYKV

Phl p 5 (SEQ ID NO: 180)MAVQKYTVALFLAVALVAGPAASYAADAGYAPATPAAAGAEAGKATTEEQKLIEDINVGFKAAVAAAASVPAADKFKTFEAAFTSSSKAATAKAPGLVPKLDAAYSVAYKAAVGATPEAKFDSFVASLTEALRVIAGALEVHAVKPVTEDPAWPKIPAGELQIIDKIDAAFKVAATAAATAPADDKFTVFEAAFNKAIKESTGGAYDTYKCIPSLEAAVKQAYAATVAAAPQVKYAVFEAALTKAITAMSEVQKVSQPATGAATVAAGAATTATGAASGAATVAAGGYKV

Phl p 5 (SEQ ID NO: 181)ADAGYAPATPAAAGAEAGKATTEEQKLIEDINVGFKAAVAAAASVPAADKFKTFEAAFTSSSKAATAKAPGLVPKLDAAYSVAYKAAVGATPEAKFDSFVASLTEALRVIAGALEVHAVKPVTEEPGMAKIPAGELQIIDKIDAAFKVAATAAATAPADDKFTVFEAAFNKAIKESTGGAYDTYKCIPSLEAAVKQAYAATVAAAPQVKYAVFEAALTKAITAMSEVQKVSQPATGAATVAAGAATTAAG AASGAATVAAGGYKV

Phl p 5 (SEQ ID NO: 182)SVKRSNGSAEVHRGAVPRRGPRGGPGRSYAADAGYAPATPAAAGAEAGKATTEEQKLIEDINVGFKAAVAAAASVPAADKFKTFEAAFTSSSKAATAKAPGLVPKLDAAYSVAYKAAVGATPEAKFDSFVASLTEALRVIAGALEVHAVKPVTEEPGMAKIPAGELQIIDKIDAAFKVAATAAATAPADDKFTVFEAAFNKAIKESTGGAYDTYKCIPSLEAAVKQAYAATVAAAPQVKYAVFEAALTKAITAMSEVQKVSQPATGAATVAAGAATTAAGAASGAATVAAGGYKV

Phl p 5 (SEQ ID NO: 183)MAVHQYTVALFLAVALVAGPAGSYAADLGYGPATPAAPAAGYTPATPAAPAGAEPAGKATTEEQKLIEKINAGFKAALAAAAGVPPADKYRTFVATFGAASNKAFAEGLSGEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATVSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDAAFKVAATAANAAPANDKFTVFEAAFNDAIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAITAMSEAQKAAKPAAAATATATAAVGAAT GAATAATGGYKV

Phl p 5 (SEQ ID NO: 175)ADLGYGGPATPAAPAEAAPAGKATTEEQKLIEKINDGFKAALAAAAGVPPADKYKTFVATFGAASNKAFAEGLSAEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDSAFKVAATAANAAPANDKFTVFEAAFNNAIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAFTAMSEAQKAAKPATEATATATAAVGAATGAATAATGGYKV

Phl p5b (SEQ ID NO: 177)AAAAVPRRGPRGGPGRSYTADAGYAPATPAAAGAAAGKATTEEQKLIEDINVGFKAAVAAAASVPAADKFKTFEAAFTSSSKAAAAKAPGLVPKLDAAYSVAYKAAVGATPEAKFDSFVASLTEALRVIAGALEVHAVKPVTEEPGMAKILQIIDKIDAAFKVAATAAATAPADDKFTVFEAAFNKAIKESTPAGEGGAYDTYKCIPSLEAAVKQAYAATVAAAPQVKYAVFEAALTKAITAMSEVQKVSQPATGAATVAAGAATTAAGAASGAATVAAGGYKV

Phl p5a (SEQ ID NO: 178)ADLGYGPATPAAPAAGYTPATPAAPAGADAAGKATTEEQKLIEKINAGFKAALAGAGVQPADKYRTFVATFGPASNKAFAEGLSGEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDAAFKVAATAANAAPANDKFTVFEAAFNDEIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAITAMSEAQKAAKPAAAATATATAAVGAATGAATAATGGYKV

Phl p 5 (SEQ ID NO: 187)AVPRRGPRGGPGRSYAADAGYAPATPAAAGAEAGKATTEEQKLIEDINVGFKAAVAAAASVPAGDKFKTFEAAFTSSSKAATAKAPGLVPKLDAAYSVAYKAAVGATPEAKFDSFVASLTEALRVIAGALEVHAVKPVTEEPGMAKIPAGELQIIDKIDAAFKVAATAAATAPADDKFTVFEAAFNKAIKESTGGAYDTYKCIPSLEAAVKQAYAATVAAAPQVKYAVFEAALTKAITAMSEVQKVSQPATGAATVAAGAATTATGAASGAATVAAGGYKV

Phl p 5b (SEQ ID NO: 188)MAVPRRGPRGGPGRSYTADAGYAPATPAAAGAAAGKATTEEQKLIEDINVGFKAAVAARQRPAADKFKTFEAASPRHPRPLRQGAGLVPKLDAAYSVAYKAAVGATPEAKFDSFVASLTEALRVIAGALEVHAVKPVTEEPGMAKIPAGELQIIDKIDAAFKVAATAAATAPADDKFTVFEAAFNKAIKESTGGAYDTYKCIPSLEAAVKQAYAATVAAAAEVKYAVFEAALTKAITAMSEVQKVSQPATGAATVAAGAATTAAGAASGAATVAAGGYKV

Phl p 5 (SEQ ID NO: 189)MAVHQYTVALFLAVALVAGPAASYAADLGYGPATPAAPAAGYTPATPAAPAEAAPAGKATTEEQKLIEKINAGFKAALAAAAGVQPADKYRTFVATFGAASNKAFAEGLSGEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDAAFKVAATAANAAPANDKFTVFEAAFNDAIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAITAMSEAQKAAKPAAAATATATAAVGAAT GAATAATGGYKV

Phl p 5 (SEQ ID NO: 190)EAPAGKATTEEQKLIEKINAGFKAALARRLQPADKYRTFVATFGPASNKAFAEGLSGEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAAELQVIEKVDAAFKVAATAANAAPANDKFTVFEAAFNDEIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAITAMSEAQKAAKPPPLPPPPQPPPLAATGAATA ATGGYKV

Phl p 5 (SEQ ID NO: 189)MAVHQYTVALFLAVALVAGPAASYAADLGYGPATPAAPAAGYTPATPAAPAEAAPAGKATTEEQKLIEKINAGFKAALAAAAGVQPADKYRTFVATFGAASNKAFAEGLSGEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDAAFKVAATAANAAPANDKFTVFEAAFNDAIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAITAMSEAQKAAKPAAAATATATAAVGAAT GAATAATGGYKV

Phl p 5b (SEQ ID NO: 188)MAVPRRGPRGGPGRSYTADAGYAPATPAAAGAAAGKATTEEQKLIEDINVGFKAAVAARQRPAADKFKTFEAASPRHPRPLRQGAGLVPKLDAAYSVAYKAAVGATPEAKFDSFVASLTEALRVIAGALEVHAVKPVTEEPGMAKIPAGELQIIDKIDAAFKVAATAAATAPADDKFTVFEAAFNKAIKESTGGAYDTYKCIPSLEAAVKQAYAATVAAAAEVKYAVFEAALTKAITAMSEVQKVSQPATGAATVAAGAATTAAGAASGAATVAAGGYKV

Phl p 5a (SEQ ID NO: 193)ADLGYGPATPAAPAAGYTPATPAAPAGADAAGKATTEEQKLIEKINAGFKAALAGAGVQPADKYRTFVATFGPASNKAFAEGLSGEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDAAFKVAATAANAAPANDKFTVFEAAFNDEIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAITAMSEAQKAAKPPPLPPPPQPPPLAATGAATAATGGYKV

Phl p 5 (SEQ ID NO: 189)MAVHQYTVALFLAVALVAGPAASYAADLGYGPATPAAPAAGYTPATPAAPAEAAPAGKATTEEQKLIEKINAGFKAALAAAAGVQPADKYRTFVATFGAASNKAFAEGLSGEPKGAAESSSKAALTSKLDAAYKLAYKTAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAAEEVKVIPAGELQVIEKVDAAFKVAATAANAAPANDKFTVFEAAFNDAIKASTGGAYESYKFIPALEAAVKQAYAATVATAPEVKYTVFETALKKAITAMSEAQKAAKPAAAATATATAAVGAAT GAATAATGGYKV

Phl p 6 (SEQ ID NO: 195)MAAHKFMVAMFLAVAVVLGLATSPTAEGGKATTEEQKLIEDVNASFRAAMATTANVPPADKYKTFEAAFTVSSKRNLADAVSKAPQLVPKLDEVYNAAYNAADHAAPEDKYEAFVLHFSEALRIIAGTPEVHAVKPGA

Phl p 6 (SEQ ID NO: 196)SKAPQLVPKLDEVYNAAYNAADHAAPEDKYEAFVLHFSEALHIIAGTPEV HAVKPGA

Phl p 6 (SEQ ID NO: 197)ADKYKTFEAAFTVSSKRNLADAVSKAPQLVPKLDEVYNAAYNAADHAAPEDKYEAFVLHFSEALHIIAGTPEVHAVKPGA

Phl p 6 (SEQ ID NO: 198)TEEQKLIEDVNASFRAAMATTANVPPADKYKTLEAAFTVSSKRNLADAVSKAPQLVPKLDEVYNAAYNAADHAAPEDKYEAFVLHFSEALRIIAGTPEVH AVKPGA

Phl p 6 (SEQ ID NO: 199)MAAHKFMVAMFLAVAVVLGLATSPTAEGGKATTEEQKLIEDINASFRAAMATTANVPPADKYKTFEAAFTVSSKRNLADAVSKAPQLVPKLDEVYNAAYNAADHAAPEDKYEAFVLHFSEALHIIAGTPEVHAVKPGA

Phl p 6 (SEQ ID NO: 200)MVAMFLAVAVVLGLATSPTAEGGKATTEEQKLIEDVNASFRAAMATTANVPPADKYKTFEAAFTVSSKRNLADAVSKAPQLVPKLDEVYNAAYNAADHAAPEDKYEAFVLHFSEALRIIAGTPEVHAVKPGA

Phl p 7 (SEQ ID NO: 201)MADDMERIFKRFDTNGDGKISLSELTDALRTLGSTSADEVQRMMAEIDTDGDGFIDFNEFISFCNANPGLMKDVAKVF

Phl p 11 (SEQ ID NO: 202)MSWQTYVDEHLMCEIEGHHLASAAILGHDGTVWAQSADFPQFKPEEITGIMKDFDEPGHLAPTGMFVAGAKYMVIQGEPGRVIRGKKGAGGITIKKTGQALVVGIYDEPMTPGQCNMVVERLGDYLVEQGMAdditional Phleum Sequences (NCBI Entrez Accession):458878; 548863; 2529314; 2529308; 2415702; 2415700; 2415698; 542168;542167; 626037; 542169; 541814; 542171; 253337; 253336; 453976; 439960.Wasp (and Related)Vespula Sequences:

465054 ALLERGEN VES V 5 (SEQ ID NO: 203)MEISGLVYLIIIVTIIDLPYGKANNYCKIKCLKGGVHTACKYGSLKPNCGNKVVVSYGLTKQEKQDILKEHNDFRQKIARGLETRGNPGPQPPAKNMKNLVWNDELAYVAQVWANQCQYGHDTCRDVAKYQVGQNVALTGSTAAKYDDPVKLVKMWEDEVKDYNPKKKFSGNDFLKTGHYTQMVWANTKEVGCGSIKYIQEKWHKHYLVCNYGPSGNFMNEELYQTK

1709545 ALLERGEN VES M 1 (SEQ ID NO: 204)GPKCPFNSDTVSIIIETRENRNRDLYTLQTLQNHPEFKKKTITRPVVFITHGFTSSASEKNFINLAKALVDKDNYMVISIDWQTAACTNEYPGLKYAYYPTAASNTRLVGQYIATITQKLVKDYKISMANIRLIGHSLGAHVSGFAGKRVQELKLGKYSEIIGLDPARPSFDSNHCSERLCETDAEYVQIIHTSNYLGTEKILGTVDFYMNNGKNNPGCGRFFSEVCSHTRAVIYMAECIKHECCLIGIPRSKSSQPISRCTKQECVCVGLNAKKYPSRGSFYVPVESTAPFCNNKGKII

1352699 ALLERGEN VES V 1 (SEQ ID NO: 205)MEENMNLKYLLLFVYFVQVLNCCYGHGDPLSYELDRGPKCPFNSDTVSIIIETRENRNRDLYTLQTLQNHPEFKKKTITRPVVFITHGFTSSASETNFINLAKALVDKDNYMVISIDWQTAACTNEAAGLKYLYYPTAARNTRLVGQYIATITQKLVKHYKISMANIRLIGHSLGAHASGFAGKKVQELKLGKYSEIIGLDPARPSFDSNHCSERLCETDAEYVQIIHTSNYLGTEKTLGTVDFYMNNGKNQPGCGRFFSEVCSHSRAVIYMAECIKHECCLIGIPKSKSSQPISSCTKQECVCVGLNAKKYPSRGSFYVPVESTAPFCNNKGKII

1346323 ALLERGEN VES V 2 (SEQ ID NO: 206)SERPKRVFNIYWNVPTFMCHQYDLYFDEVTNFNIKRNSKDDFQGDKIAIFYDPGEFPALLSLKDGKYKKRNGGVPQEGNITIHLQKFIENLDKIYPNRNFSGIGVIDFERWRPIFRQNWGNMKIHKNFSIDLVRNEHPTWNKKMIELEASKRFEKYARFFMEETLKLAKKTRKQADWGYYGYPYCFNMSPNNLVPECDVTAMHENDKMSWLFNNQNVLLPSVYVRQELTPDQRIGLVQGRVKEAVRISNNLKHSPKVLSYWWYVYQDETNTFLTETDVKKTFQEIVINGGDGIIIWGSSSDVNSLSKCKRLQDYLLTVLGPIAINVTEAVN

549194 ALLERGEN VES VI (SEQ ID NO: 207)5KVNYCKIKCLKGGVHTACKYGTSTKPNCGKMVVKAYGLTEAEKQEILKVHNDFRQKVAKGLETRGNPGPQPPAKNMNNLVWNDELANIAQVWASQCNYGHDTCKDTEKYPVGQNIAKRSTTAALFDSPGKLVKMWENEVKDFNPNIEWSKNNLKKTGHYTQMVWAKTKEIGCGSVKYVKDEWYTHYLVCNYGPSGNFR NEKLYEKKAdditional Vespula Sequences (NCBI Entrez Accession):549193; 549192; 549191; 549190; 5491104; 117414; 126761; 69576; 625255;6271104; 627188; 627187; 482382; 112561; 627186; 627185; 1923233;1047645; 1047647; 745570; 225764; 162551.Tree Allergen Sequences (Mainly Birch) Sequences:

114922 Bet v 1 (SEQ ID NO: 208)MGVFNYETETTSVIPAARLFKAFILDGDNLFPKVAPQAISSVENIEGNGGPGTIKKISFPEGFPFKYVKDRVDEVDHTNFKYNYSVIEGGPIGDTLEKISNEIKIVATPDGGSILKISNKYHTKGDHEVKAEQVKASKEMGETLLRAVESYLL AHSDAYN

130975 Bet v 2 (SEQ ID NO: 209)MSWQTYVDEHLMCDIDGQASNSLASAIVGHDGSVWAQSSSFPQFKPQEITGIMKDFEEPGHLAPTGLHLGGIKYMVIQGEAGAVIRGKKGSGGITIKKTGQALVFGIYEEPVTPGQCNMVVERLGDYLIDQGL

1168696 Bet v 3 (SEQ ID NO: 210)MPCSTEAMEKAGHGHASTPRKRSLSNSSFRLRSESLNTLRLRRIFDLFDKNSDGIITVDELSRALNLLGLETDLSELESTVKSFTREGNIGLQFEDFISLHQSLNDSYFAYGGEDEDDNEEDMRKSILSQEEADSFGGFKVFDEDGDGYISARELQMVLGKLGFSEGSEIDRVEKMIVSVDSNRDGRVDFFEFKDMMRSVLVRS S

809536 Bet v 4 (SEQ ID NO: 211)MADDHPQDKAERERIFKRFDANGDGKISAAELGEALKTLGSITPDEVKHNIMAEIDTDGDGFISFQEFTDFGRANRGLLKDVAKIF

543675 Que a I-Quercus alba = oak trees (fragment) (SEQ ID NO: 212)GVFTXESQETSVIAPAXLFKALFL

543509 Car b I-Carpinus betulus = hornbeam trees (fragment)(SEQ ID NO: 213) GVFNYEAETPSVIPAARLFKSYVLDGDKLIPKVAPQAIXK

543491 Aln g I-Alnus glutinosa = alder trees (fragment) (SEQ ID NO: 214)GVFNYEAETPSVIPAARLFKAFILDGDKLLPKVAPEAVSSVENI

1204056 Rubisco (SEQ ID NO: 215)VQCMQVWPPLGLKKFETLSYLPPLSSEQLAKEVDYLLRKNLIPCLEFELEHGFVYREHNRSPGYYDGRYWTMWKLPMFGCNDSSQVLKELEECKKAYPSAFI RIIGFDDKAdditional Tree Allergen Sequences (NCBI Entrez Accession Number):131919; 128193; 585564; 1942360; 2554672; 2392209; 2414158; 1321728;1321726; 1321724; 1321722; 1321720; 1321718; 1321716; 1321714; 1321712;3015520; 2935416; 464576; 1705843; 1168701; 1168710; 1168709; 1168708;1168707; 1168706; 1168705; 1168704; 1168703; 1168702; 1842188; 2564228;2564226; 2564224; 2564222; 2564220; 2051993; 18131041; 15368104; 534910;534900; 5341048; 1340000; 1339998; 2149808; 66207; 2129477; 1076249;1076247; 629480; 481805; 81443; 1361968; 1361967; 1361966; 1361965;1361964; 1361963; 1361962; 1361961; 1361960; 1361959; 320546; 629483;629482; 629481; 541804; 320545; 81444; 541814; 629484; 474911; 452742;1834387; 298737; 298736; 1584322; 1584321; 584320; 1542873; 1542871;1542869; 1542867; 1542865; 1542863; 1542861; 1542859; 1542857; 1483232;1483230; 1483228; 558561; 551640; 488605; 452746; 452744; 452740;452738; 452736; 452734; 452732; 452730; 452728; 450885; 17938; 17927;17925; 17921; 297538; 510951; 2104331; 2104329; 166953.PeanutPeanut Sequences

1168391 Ara h 1 (SEQ ID NO: 216)MRGRVSPLMLLLGILVLASVSATHAKSSPYQKKTENPCAQRCLQSCQQEPDDLKQKACESRCTKLEYDPRCVYDPRGHTGTTNQRSPPGERTRGRQPGDYDDDRRQPRREEGGRWGPAGPREREREEDWRQPREDWRRPSHQQPRKIRPEGREGEQEWGTPGSHVREETSRNNPFYFPSRRFSTRYGNQNGRIRVLQRFDQRSRQFQNLQNHRIVQIEAKPNTLVLPKHADADNILVIQQGQATVTVANGNNRKSFNLDEGHALRIPSGFISYILNRHDNQNLRVAKISMPVNTPGQFEDFFPASSRDQSSYLQGFSRNTLEAAFNAEFNEIRRVLLEENAGGEQEERGQRRWSTRSSENNEGVIVKVSKEHVEELTKHAKSVSKKGSEEEGDITNPINLREGEPDLSNNFGKLFEVKPDKKNPQLQDLDMMLTCVEIKEGALMLPHFNSKAMVIVVVNKGTGNLELVAVRKEQQQRGRREEEEDEDEEEEGSNREVRRYTARLKEGDVFIMPAAHPVAINASSELHLLGFGINAENNHRIFLAGDKDNVIDQIEKQAKDLAFPGSGEQVEKLIKNQKESHFVSARPQSQSQSPSSPEKESPEKEDQEEENQ GGKGPLLSILKAFNRagweedAmbrosia Sequences

113478 Amb a 1 (SEQ ID NO: 217)MGIKHCCYILYFTLALVTLLQPVRSAEDLQQILPSANETRSLTTCGTYNIIDGCWRGKADWAENRKALADCAQGFAKGTIGGKDGDIYTVTSELDDDVANPKEGTLRFGAAQNRPLWIIFARDMVIRLDRELAINNDKTIDGRGAKVEIINAGFAIYNVKNIIIHNIIMHDIVVNPGGLIKSHDGPPVPRKGSDGDAIGISGGSQIWIDHCSLSKAVDGLIDAKHGSTHFTVSNCLFTQHQYLLLFWDFDERGMLCTVAFNKFTDNVDQRMPNLRHGFVQVVNNNYERWGSYALGGSAGPTILSQGNRFLASDIKKEVVGRYGESAMSESMWNWRSYMDVFENGAIFVPSGVDPVLTPEQNAGMIPAEPGEAVLRLTSSAGVLSCQPGAPC

113479 Amb a 2 (SEQ ID NO: 218)MGIKHCCYILYFTLALVTLVQAGRLGEEVDILPSPNDTRRSLQGCEAHNIIDKCWRCKPDWAENRQALGNCAQGFGKATHGGKWGDIYMVTSDQDDDVVNPKEGTLRFGATQDRPLWIIFQRDMIIYLQQEMVVTSDKTIDGRGAKVELVYGGITLMNVKNVIIHNIDIHDVRVLPGGRIKSNGGPAIPRHQSDGDAIHVTGSSDIWIDHCTLSKSFDGLVDVNWGSTGVTISNCKFTHHEKAVLLGASDTHFQDLKMHVTLAYNIFTNTVHERMPRCRFGFFQIVNNFYDRWDKYAIGGSSNPTILSQGNKFVAPDFIYKKNVCLRTGAQEPEWMTWNWRTQNDVLENGAIFVASGSDPVLTAEQNAGMMQAEPGDMVPQLTMNAGVLTCSPGAPC

113477 Amb a 1.3 (SEQ ID NO: 219)MGIKQCCYILYFTLALVALLQPVRSAEGVGEILPSVNETRSLQACEALNIIDKCWRGKADWENNRQALADCAQGFAKGTYGGKWGDVYTVTSNLDDDVANPKEGTLRFAAAQNRPLWIIFKNDMVINLNQELVVNSDKTIDGRGVKVEIINGGLTLMNVKNIIIHNINIHDVKVLPGGMIKSNDGPPILRQASDGDTINVAGSSQIWIDHCSLSKSFDGLVDVTLGSTHVTISNCKFTQQSKAILLGADDTHVQDKGMLATVAFNMFTDNVDQRMPRCRFGFFQVVNNNYDRWGTYAIGGSSAPTILCQGNRFLAPDDQIKKNVLARTGTGAAESMAWNWRSDKDLLENGAIFVTSGSDPVLTPVQSAGMIPAEPGEAAIKLTSSAGVFSCHPGAPC

113476 Amb a 1.2 (SEQ ID NO: 220)MGIKHCCYILYFTLALVTLLQPVRSAEDVEEFLPSANETRRSLKACEAHNIIDKCWRCKADWANNRQALADCAQGFAKGTYGGKHGDVYTVTSDKDDDVANPKEGTLRFAAAQNRPLWIIFKRNIVIVIHLNQELVVNSDKTIDGRGVKVNIVNAGLTLMNVKNIIIHNINIHDIKVCPGGMIKSNDGPPILRQQSDGDAINVAGSSQIWIDHCSLSKASDGLLDITLGSSHVTVSNCKFTQHQFVLLLGADDTHYQDKGMLATVAFNMFTDHVDQRMPRCRFGFFQVVNNNYDRWGTYAIGGSSAPTILSQGNRFFAPDDIIKKNVLARTGTGNAESMSWNWRTDRDLLENGAIFLPSGSDPVLTPEQKAGMIPAEPGEAVLRLTSSAGVLSCHQGAPC

113475 Amb a 1.1 (SEQ ID NO: 221)MGIKHCCYILYFTLALVTLLQPVRSAEDLQEILPVNETRRLTTSGAYNIIDGCWRGKADWAENRKALADCAQGFGKGTVGGKDGDIYTVTSELDDDVANPKEGTLRFGAAQNRPLWIIFERDMVIRLDKEMVVNSDKTIDGRGAKVEIINAGFTLNGVKNVIIHNINMHDVKVNPGGLIKSNDGPAAPRAGSDGDAISISGSSQIWIDHCSLSKSVDGLVDAKLGTTRLTVSNSLFTQHQFVLLFGAGDENIEDRGMLATVAFNTFTDNVDQRMPRCRHGFFQVVNNNYDKWGSYAIGGSASPTILSQGNRFCAPDERSKKNVLGRHGEAAAESMKWNWRTNKDVLENGAIFVASGVDPVLTPEQSAGMIPAEPGESALSLTSSAGVLSCQPGAPCCedar Sequences

493634 Cry j IB precursor (SEQ ID NO: 222)MDSPCLVALLVFSFVIGSCFSDNPIDSCWRGDSNWAQNRMKLADCAVGFGSSTMGGKGGDLYTVTNSDDDPVNPPGTLRYGATRDRPLWIIFSGNMNIKLKMPMYIAGYKTFDGRGAQVYIGNGGPCVFIKRVSNVIIHGLYLYGCSTSVLGNVLINESFGVEPVHPQDGDALTLRTATNIWIDHNSFSNSSDGLVDVTLTSTGVTISNNLFFNHHKVMSLGHDDAYSDDKSMKVTVAFNQFGPNCGQRMPRARYGLVHVANNNYDPWTIYAIGGSSNPTILSEGNSFTAPNESYKKQVTIRIGCKTSSSCSNWVWQSTQDVFYNGAYFVSSGKYEGGNIYTKKEAFNVENGNATPHLTQNAGVLTCSLSKRC

493632 Cry j IA precursor (SEQ ID NO: 223)MDSPCLVALLVLSFVIGSCFSDNPIDSCWRGDSNWAQNRMKLADCAVGFGSSTMGGKGGDLYTVTNSDDDPVNPAPGTLRYGATRDRPLWIIFSGNMNIKLKMPMYIAGYKTFDGRGAQVYIGNGGPCVFIKRVSNVIIHGLHLYGCSTSVLGNVLINESFGVEPVHPQDGDALTLRTATNIWIDHNSFSNSSDGLVDVTLSSTGVTISNNLFFNHHKVMLLGHDDAYSDDKSMKVTVAFNQFGPNCGQRMPRARYGLVHVANNNYDPWTIYAIGGSSNPTILSEGNSFTAPNESYKKQVTIRIGCKTSSSCSNWVWQSTQDVFYNGAYFVSSGKYEGGNIYTKKEAFNVENGNATPQLTKNAGVLTCSLSKRC

1076242 Cry j II precursor - Japanese cedar (SEQ ID NO: 224)MAMKLIAPMAFLAMQLIIMAAAEDQSAQIMLDSVVEKYLRSNRSLRKVEHSRHDAINIFNVEKYGAVGDGKHDCTEAFSTAWQAACKNPSAMLLVPGSKKFVVNNLFFNGPCQPHFTFKVDGIIAAYQNPASWKNNRIWLQFAKLTGFTLMGKGVIDGQGKQWWAGQCKWVNGREICNDRDRPTAIKFDFSTGLIIQGLKLMNSPEFHLVFGNCEGVKIIGISITAPRDSPNTDGIDIFASKNFHLQKNTIGTGDDCVAIGTGSSNIVIEDLICGPGHGISIGSLGRENSRAEVSYVHVNGAKFIDTQNGLRIKTWQGGSGMASHIIYENVEMINSENPILINQFYCTSASACQNQRSAVQIQDVTYKNIRGTSATAAAIQLKCSDSMPCKDIKLSDISLKLTSGKIASCLNDNANGYFSGHVIPACKNLSPSAKRKESKSHKHPKTVMVENMRAYDKGNRTRILLGSRPPNCTNKCHGCSPCKAKLVIVHRIMPQEYYP QRWICSCHGKIYHP

1076241 Cry j II protein - Japanese cedar (SEQ ID NO: 225)MAMKFIAPMAFVAMQLIIIVIAAAEDQSAQIMLDSDIEQYLRSNRSLRKVEHSRHDAINIFNVEKYGAVGDGKHDCTEAFSTAWQAACKKPSAMLLVPGNKKFVVNNLFFNGPCQPHFTFKVDGIIAAYQNPASWKNNRIWLQFAKLTGFTLMGKGVIDGQGKQWWAGQCKWVNGREICNDRDRPTAIKFDFSTGLIIQGLKLMNSPEFHLVFGNCEGVKIIGISITAPRDSPNTDGIDIFASKNFHLQKNTIGTGDDCVAIGTGSSNIVIEDLICGPGHGISIGSLGRENSRAEVSYVHVNGAKFIDTQNGLRIKTWQGGSGMASHIIYENVEMINSENPILINQFYCTSASACQNQRSAVQIQDVTYKNIRGTSATAAAIQLKCSDSMPCKDIKLSDISLKLTSGKIASCLNDNANGYFSGHVIPACKNLSPSAKRKESKSHKHPKTVMVKNMGAYDKGNRTRILLGSRPPNCTNKCHGCSPCKAKLVIVHRIMPQEY YPQRWMCSRHGKIYHP

541803 Cry j I precursor - Japanese cedar (SEQ ID NO: 226)MDSPCLVALLVLSFVIGSCFSDNPIDSCWRGDSNWAQNRMKLADCAVGFGSSTMGGKGGDLYTVTNSDDDPVNPPGTLRYGATRDRPLWIIFSGNMNIKLKMPMYIAGYKTFDGRGAQVYIGNGGPCVFIKRVSNVIIHGLHLYGCSTSVLGNVLINESFGVEPVHPQDGDALTLRTATNIWIDHNSFSNSSDGLVDVTLSSTGVTISNNLFFNHHKVMLLGHDDAYSDDKSMKVTVAFNQFGPNCGQRMPRARYGLVHVANNNYDPWTIYAIGGSSNPTILSEGNSFTAPNESYKKQVTIRIGCKTSSSCSNWVWQSTQDVFYNGAYFVSSGKYEGGNIYTKKEAFNVENGNATPQLTKNAGVLTCSLSKRC

541802 Cry j I precursor- Japanese cedar (SEQ ID NO: 227)MDSPCLVALLVFSFVIGSCFSDNPIDSCWRGDSNWAQNRMKLADCAVGFGSSTMGGKGGDLYTVTNSDDDPVNPAPGTLRYGATRDRPLWIIFSGNMNIKLKMPMYIAGYKTFDGRGAQVYIGNGGPCVFIKRVSNVIIHGLYLYGCSTSVLGNVLINESFGVEPVHPQDGDALTLRTATNIWIDHNSFSNSSDGLVDVTLTSTGVTISNNLFFNHHKVMSLGHDDAYSDDKSMKVTVAFNQFGPNCGQRMPRARYGLVHVANNNYDPWTIYAIGGSSNPTILSEGNSFTAPNESYKKQVTIRIGCKTSSSCSNWVWQSTQDVFYNGAYFVSSGKYEGGNIYTKKEAFNVENGNATPHLTQNAGVLTCSLSKRCDogCanis Sequences:

Can f 1 (SEQ ID NO: 228)MKTLLLTIGFSLIAILQAQDTPALGKDTVAVSGKWYLKAMTADQEVPEKPDSVTPMILKAQKGGNLEAKITMLTNGQCQNITVVLHKTSEPGKYTAYEGQRVVFIQPSPVRDHYILYCEGELHGRQIRMAKLLGRDPEQSQEALEDFREFSRAKGLNQEILELAQSETCSPGGQ

Serum albumin fragment (SEQ ID NO: 229) EAYKSEIAHRYNDLGEEHFRGLVL

Serum albumin fragment (SEQ ID NO: 230)LSSAKERFKCASLQKFGDRAFKAWSVARLSQRFPKADFAEISKVVTDLTKVHKECCHGDLLECADDRADLAKYMCENQDSISTKLKECCDKPVLEKSQCLAEVERDELPGDLPSLAADFVEDKEVCKNYQEAKDVFLGTFLYEYSRRHPEYSVSLLLRLAKEYEATLEKCCATDDPPTCYAKVLDEFKPLVDEPQNLVKTNCELFEKLGEYGFQNALLVRYTKKAPQVSTPTLVVEVSRKLGKVGTKCCK KPESERMSCADDFLS

Can f 2 (SEQ ID NO: 231)MQLLLLTVGLALICGLQAQEGNHEEPQGGLEELSGRWHSVALASNKSDLIKPWGHFRVFIHSMSAKDGNLHGDILIPQDGQCEKVSLTAFKTATSNKFDLEYWGHNDLYLAEVDPKSYLILYMINQYNDDTSLVAHLMVRDLSRQQDFLPAFESVCEDIGLHKDQIVVLSDDDRCQGSRDAdditional Dog Allergen Protein (NCBI Entrez Accession):1731859HorseEquus Sequences:

1575778 Equ c1 (SEQ ID NO: 232)MKLLLLCLGLILVCAQQEENSDVAIRNFDISKISGEWYSIFLASDVKEKIEENGSMRVFVDVIRALDNSSLYAEYQTKVNGECTEFPMVFDKTEEDGVYSLNYDGYNVFRISEFENDEHIILYLVNFDKDRPFQLFEFYAREPDVSPEIKEEFVKIVQKRGIVKENIIDLTKIDRCFQLRGNGVAQA

3121755 Equ c 2 (SEQ ID NO: 233) SQXPQSETDYSQLSGEWNTIYGAASNIXKEuroglyphus (Mite)Euroglyphus Sequences:

Eur m 1 (variant) (SEQ ID NO: 234)TYACSINSVSLPSELDLRSLRTVTPIRMQGGCGSCWAFSGVASTESAYLAYRNMSLDLAEQELVDCASQNGCHGDTIPRGIEYIQQNGVVQEHYYPYVAREQSCHRPNAQRYGLKNYCQISPPDSNKIRQALTQTHTAVAVIIGIKDLNAFRHYDGRTIMQHDNGYQPNYHAVNIVGYGNTQGVDYWIVRNSWDTTWGDN GYGYFAANINL

Eur m 1 (variant) (SEQ ID NO: 234)TYACSINSVSLPSELDLRSLRTVTPIRMQGGCGSCWAFSGVASTESAYLAYRNMSLDLAEQELVDCASQNGCHGDTIPRGIEYIQQNGVVQEHYYPYVAREQSCHRPNAQRYGLKNYCQISPPDSNKIRQALTQTHTAVAVIIGIKDLNAFRHYDGRTIMQHDNGYQPNYHAVNIVGYGNTQGVDYWIVRNSWDTTWGDN GYGYFAANINL

Eur m 1 (variant) (SEQ ID NO: 236)ETNACSINGNAPAEIDLRQMRTVTPIRMQGGCGSCWAFSGVAATESAYLAYRNQSLDLAEQELVDCASQHGCHGDTIPRGIEYIQHNGVVQESYYRYVAREQSCRRPNAQRFGISNYCQIYPPNANKIREALAQTHSAIAVIIGIKDLDAFRHYDGRTIIQRDNGYQPNYHAVNIVGYSNAQGVDYWIVRNSWDTNWGDN GYGYFAANIDL

Eur m 1 (variant) (SEQ ID NO: 237)ETSACRINSVNVPSELDLRSLRTVTPIRMQGGCGSCWAFSGVAATESAYLAYRNTSLDLSEQELVDCASQHGCHGDTIPRGIEYIQQNGVVEERSYPYVAREQQCRRPNSQHYGISNYCQIYPPDVKQIREALTQTHTAIAVIIGIKDLRAFQHYDGRTIIQHDNGYQPNYHAVNIVGYGSTQGVDYWIVRNSWDTTWGDSGYGYFQA GNNLPoa (Grass) Sequences

113562 POLLEN ALLERGEN POA P 9 (SEQ ID NO: 238)MAVQKYTVALFLVALVVGPAASYAADLSYGAPATPAAPAAGYTPAAPAGAAPKATTDEQKMIEKINVGFKAAVAAAGGVPAANKYKTFVATFGAASNKAFAEALSTEPKGAAVDSSKAALTSKLDAAYKLAYKSAEGATPEAKYDDYVATLSEALRIIAGTLEVHGVKPAAEEVKATPAGELQVIDKVDAAFKVAATAANAAPANDKFTVFEAAFNDAIKASTGGAYQSYKFIPALEAAVKQSYAATVATAPAVKYTVFETALKKAITAMSQAQKAAKPAAAATGTATAAVGAATGAATAAAGG YKV

113561 POA P 9 (SEQ ID NO: 239)MAVHQYTVALFLAVALVAGPAASYAADVGYGAPATLATPATPAAPAAGYTPAAPAGAAPKATTDEQKLIEKINAGFKAAVAAAAGVPAVDKYKTFVATFGTASNKAFAEALSTEPKGAAAASSNAVLTSKLDAAYKLAYKSAEGATPEAKYDAYVATLSEALRIIAGTLEVHAVKPAGEEVKAIPAGELQVIDKVDAAFKVAATAANAAPANDKFTVFEAAFNDAIKASTGGAYQSYKFIPALEAAVKQSYAATVATAPAVKYTVFETALKKAITAMSQAQKAAKPAAAVTATATGAVGAATGAVGAATGAATAAAGGYKTGAATPTAGGYKV

113560 POA P 9 (SEQ ID NO: 240)MDKANGAYKTALKAASAVAPAEKFPVFQATFDKNLKEGLSGPDAVGFAKKLDAFIQTSYLSTKAAEPKEKFDLFVLSLTEVLRFMAGAVKAPPASKFPAKPAPKVAAYTPAAPAGAAPKATTDEQKLIEKINVGFKAAVAAAAGVPAASKYKTFVATFGAASNKAFAEALSTEPKGAAVASSKAVLTSKLDAAYKLAYKSAEGATPEAKYDAYVATLSEALRIIAGTLEVHGVKPAAEEVKAIPAGELQVIDKVDAAFKVAATAANAAPANDKFTVFEAAFNDAIKASTGGAYQSYKFIPALEAAVKQSYAATVATAPAVKYTVFETALKKAITAMSQAQKAAKPAAAVTGTATSAVG AATGAATAAAGGYKVCockroach Sequences

2833325 Cr p1 (SEQ ID NO: 241)MKTALVFAAVVAFVAARFPDHKDYKQLADKQFLAKQRDVLRLFHRVHQHNILNDQVEVGIPMTSKQTSATTVPPSGEAVHGVLQEGHARPRGEPFSVNYEKHREQAIMLYDLLYFANDYDTFYKTACWARDRVNEGMFMYSFSIAVFHRDDMQGVMLPPPYEVYPYLFVDHDVIHMAQKYWMKNAGSGEHHSHVIPVNFTLRTQDHLLAYFTSDVNLNAFNTYYRYYYPSWYNTTLYGHNIDRRGEQFYYTYKQIYARYFLERLSNDLPDVYPFYYSKPVKSAYNPNLRYHNGEEMPVRPSNMYVTNFDLYYIADIKNYEKRVEDAIDFGYAFDEHMKPHSLYHDVHGMEYLADMIEGNMDSPNFYFYGSIYHMYHSMIGHIVDPYHKMGLAPSLEHPETVLRDPVFYQLWKRVDHLFQKYKNRLPRYTHDELAFEGVKVENVDVGKLYTYFEQYDMSLDMAVYVNNVDQISNVDVQLAVRLNHKPFTYNIEVSSDKAQDVYVAVFLGPKYDYLGREYDLNDRRHYFVEMDRFPYHVGAGKTVIERNSHDSNIIAPERDSYRTFYKKVQEAYEGKSQYYVDKGHNYCGYPENLLIPKGKKGGQAYTFYVIVTPYVKQDEHDFEPYNYKAFSYCGVGSERKYPDNKPLGYPFDRKIYSNDFYTPNMYFKDVIIFHKKYDEVGVQGH

2231297 Cr p2 (SEQ ID NO: 242)INEIHSIIGLPPFVPPSRRHARRGVGINGLIDDVIAILPVDELKALFQEKL- ETSPDFKALYDAIRSPEFQSITSTLNAMQRSEHHQNLRDKGVDVDHFIQLIRAL- FGLSRAARNLQDDLNDFLHSLEPTSPRHRHGLPRQRRRSARVSAYLHADDFHK- IITTIEALPEFANFYNFLKEHGLDVVDYINEIHSIIGLPPFVPPSRRHARRGV- GINGLIDDVIAILPVDELKALFQEKLETSPDFKALYDAIRSPEFQSIISTLNA- MPEYQELLQNLRDKGVDVDHFIRVDQGTLRTLSSGQRNLQDDLNDFLALIPTD- QILAIAMDYLANDAEVQELVAYLQSDDFHKIITTIEALPEFANFYNFLKEHGL- DVVDYINEIHSIIGLPPFVPPSQRHARRGVGINGLIDDVIAILPVDELKALFQ- EKLETSPDF KALYDAIDLRSSRA

1703445 Bla g 2 (SEQ ID NO: 243)MIGLKLVTVLFAVATITHAAELQRVPLYKLVHVFINTQYAGITKIGNQNFLTVFDSTSCNVVVASQECVGGACVCPNLQKYEKLKPKYISDGNVQVKFFDTGSAVGRGIEDSLTISNLTTSQQDIVLADELSQEVCILSADVVVGIAAPGCPNALKGKTVLENFVEENLIAPVFSIHHARFQDGEHFGEBFGGSDWKYVDGEFTYVPLVGDDSWKFRLDGVKIGDTTVAPAGTQMIDTSKAIIVGPKAYVNPINEAIGCVVEKTTTRRICKLDCSKIPSLPDVTFVINGRNFNISSQYYIQQNGNLCYSGFQPCGHSDHFFIGDFFVDHYYSEFNWENKTMGFGRSVE SV

1705483 Bla g 4 (SEQ ID NO: 244)AVLALCATDTLANEDCFRHESLVPNLDYERFRGSWIIAAGTSEALTQYKCWIDRFSYDDALVSKYTDSQGKNRTTIRGRTKFEGNKFTIDYNDKGKAFSAPYSVLATDYENYAIVEGCPAAANGHVIYVQIRFSVRRFHPKLGDKEMIQHYTLDQVNQHKKAIEEDLKHFNLKYEDLHSTCH

2326190 Bla g 5 (SEQ ID NO: 245)YKLTYCPVKALGEPIRFLLSYGEKDFEDYRFQEGDWPNLKPSMPFGKTPVLEIDGKQTHQSVAISRYLGKQFGLSGKDDWENLEIDMIVDTISDFRAAIANYHYDADENSKQKKWDPLKKETIPYYTKKFDEVVKANGGYLAAGKLTWADFYFVAILDYLNHMAKEDLVANQPNLKALREKVLGLPAIKAWVAKRPPTDLAdditional cockroach sequences (NCBI Entrez accession numbers):2580504; 1580797; 1580794; 1362590; 544619; 544618; 15315104; 1580792;1166573; 1176397; 21047849.Allergen (General) Sequences:NCBI Accession Numbers 2739154; 3719257; 3703107; 3687326; 3643813;3087805; 1864024; 1493836; 1480457; 25910476; 25910474; 1575778; 763532;746485; 163827; 163823; 3080761; 163825; 3608493; 3581965; 2253610;2231297; 21047849; 3409499; 3409498; 3409497; 3409496; 3409495; 3409494;3409493; 3409492; 3409491; 3409490; 34094104; 3409488; 3409487; 3409486;3409485; 3409484; 3409483; 3409482; 3409481; 3409480; 3409479; 3409478;3409477; 3409476; 3409475; 3409474; 3409473; 3409472; 3409471; 3409470;3409469; 3409468; 3409467; 3409466; 3409465; 3409464; 3409463; 3409462;3409461; 3409460; 3409459; 3409458; 3409457; 3409456; 3318885; 3396070;3367732; 1916805; 3337403; 2851457; 2851456; 1351295; 549187; 136467;1173367; 2499810; 2498582; 2498581; 1346478; 1171009; 126608; 114091;2506771; 1706660; 1169665; 1169531; 232086; 4161048; 114922; 2497701;1703232; 1703233; 1703233; 1703232; 3287877; 3122132; 3182907; 3121758;3121756; 3121755; 3121746; 3121745; 3319925; 3319923; 3319921; 3319651;33187104; 3318779; 3309647; 3309047; 3309045; 3309043; 3309041; 3309039;3288200; 3288068; 2924494; 3256212; 3256210; 3243234; 3210053; 3210052;3210051; 3210050; 3210049; 3210048; 3210047; 3210046; 3210045; 3210044;3210043; 3210042; 3210041; 3210040; 3210039; 3210038; 3210037; 3210036;3210035; 3210034; 3210033; 3210032; 3210031; 3210030; 3210029; 3210028;3210027; 3210026; 3210025; 3210024; 3210023; 3210022; 3210021; 3210020;3210019; 3210018; 3210017; 3210016; 3210015; 3210014; 3210013; 3210012;3210011; 3210010; 3210009; 3210008; 3210007; 3210006; 3210005; 3210004;3210003; 3210002; 3210001; 3210000; 3209999; 3201547; 2781152; 2392605;2392604; 2781014; 1942360; 2554672; 2392209; 3114481; 3114480; 2981657;3183706; 3152922; 3135503; 3135501; 3135499; 3135497; 2414158; 1321733;1321731; 1321728; 1321726; 1321724; 1321722; 1321720; 1321718; 1321716;1321714; 1321712; 3095075; 3062795; 3062793; 3062791; 2266625; 2266623;2182106; 3044216; 2154736; 3021324; 3004467; 3005841; 3005839; 3004485;3004473; 3004471; 3004469; 3004465; 2440053; 1805730; 2970629; 29591048;2935527; 2935416; 809536; 730091; 585279; 584968; 2498195; 2833325;2498604; 2498317; 2498299; 2493414; 2498586; 2498585; 2498576; 2497749;2493446; 2493445; 1513216; 729944; 2498099; 548449; 465054; 465053;465052; 548671; 548670; 548660; 548658; 548657; 2832430; 232084;2500822; 2498118; 2498119; 2498119; 2498118; 1708296; 1708793; 416607;416608; 416608; 416607; 2499791; 2498580; 2498579; 2498578; 2498577;2497750; 1705483; 1703445; 1709542; 1709545; 17105104; 1352699; 1346568;1346323; 1346322; 2507248; 11352240; 1352239; 1352237; 1352229; 1351935;1350779; 1346806; 1346804; 1346803; 1170095; 1168701; 1352506; 1171011;1171008; 1171005; 1171004; 1171002; 1171001; 1168710; 1168709; 1168708;1168707; 1168706; 1168705; 1168704; 1168703; 1168702; 1168696; 1168391;1168390; 1168348; 1173075; 1173074; 1173071; 1169290; 11610470; 1168402;729764; 729320; 729979; 729970; 729315; 730050; 730049; 730048; 549194;549193; 549192; 549191; 549190; 5491104; 549188; 549185; 549184; 549183;549182; 549181; 549180; 549179; 464471; 585290; 416731; 1169666; 113478;113479; 113477; 113476; 113475; 130975; 119656; 113562; 113561; 113560;416610; 126387; 126386; 126385; 132270; 416611; 416612; 416612; 416611;730035; 127205; 1352238; 125887; 549186; 137395; 730036; 133174; 114090;131112; 126949; 129293; 124757; 129501; 416636; 2801531; 2796177;2796175; 2677826; 2735118; 2735116; 2735114; 2735112; 2735110; 2735108;2735106; 2735104; 2735102; 2735100; 2735098; 2735096; 2707295; 2154730;2154728; 1684720; 2580504; 2465137; 2465135; 2465133; 2465131; 2465129;2465127; 2564228; 2564226; 2564224; 2564222; 2564220; 2051993; 1313972;1313970; 1313968; 1313966; 2443824; 2488684; 2488683; 2488682; 2488681;2488680; 2488679; 2488678; 2326190; 2464905; 2415702; 2415700; 2415698;2398759; 2398757; 2353266; 2338288; 1167836; 414703; 2276458; 1684718;2293571; 1580797; 1580794; 2245508; 2245060; 1261972; 2190552; 1881574;511953; 1532058; 1532056; 1532054; 1359436; 666007; 487661; 217308;1731859; 217306; 217304; 1545803; 1514943; 577696; 516728; 506858;493634; 493632; 2154734; 2154732; 543659; 1086046; 1086045; 2147643;2147642; 1086003; 1086002; 1086001; 543675; 543623; 543509; 543491;1364099; 2147108; 2147107; 1364001; 1085628; 631913; 631912; 631911;2147092; 477301; 543482; 345521; 542131; 542130; 542129; 100636;2146809; 480443; 2114497; 2144915; 72355; 71728; 319828; 1082946;1082945; 1082944; 539716; 539715; 423193; 423192; 423191; 423190;1079187; 627190; 6271104; 627188; 627187; 482382; 1362656; 627186;627185; 627182; 482381; 85299; 85298; 2133756; 2133755; 1079186; 627181;321044; 321043; 112559; 112558; 1362590; 2133564; 1085122; 10710471;627144; 627143; 627142; 627141; 280576; 102835; 102834; 102833; 102832;84703; 84702; 84700; 84699; 84698; 84696; 477888; 477505; 102575;102572; 478272; 2130094; 629813; 629812; 542172; 542168; 542167; 481432;320620; 280414; 626029; 542132; 320615; 320614; 100638; 100637; 100635;82449; 320611; 320610; 280409; 320607; 320606; 539051; 539050; 539049;539048; 322803; 280407; 100501; 100498; 100497; 100496; 1362137;1362136; 1362135; 1362134; 1362133; 1362132; 1362131; 1362130; 1362129;1362128; 100478; 21291041; 1076531; 1362049; 1076486; 2129817; 2129816;2129815; 2129814; 2129813; 2129812; 2129805; 2129804; 2129802; 2129801;2129800; 2129799; 479902; 479901; 2129477; 1076247; 629480; 1076242;1076241; 541803; 541802; 280372; 280371; 1361968; 1361967; 1361966;1361965; 1361964; 1361963; 1361962; 1361961; 1361960; 1361959; 320546;2119763; 543622; 541804; 478825; 478824; 478823; 421788; 320545; 81444;626037; 626028; 539056; 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1247377; 1247375; 1247373; 862307; 312284;999462; 999460; 999458; 587450; 763064; 886209; 1176397; 1173557;902012; 997915; 997914; 997913; 997912; 997911; 997910; 99790; 997908;997907; 997906; 997905; 997904; 997903; 997902; 997901; 997900; 9971049;9971048; 9971047; 9971046; 9971045; 9971044; 9971043; 9971042; 910984;910983; 910982; 910981; 511604; 169631; 169629; 169627; 168316; 168314;607633; 555616; 293902; 485371; 455288; 166447; 166445; 166443; 166435;162551; 160780; 552080; 156719; 156715; 515957; 515956; 515955; 515954;515953; 459163; 166953; 386678; 169865.Particularly preferred allergens/antigens include: cat dander proteinFel dl; House dust mite proteins Der P1, Der P2 and Der P7; Ragweedprotein amb a 1.1, a 1.2, a 1.3 or a 1.4; Rye grass proteins lol p1 andlol p5; Timothy grass proteins phl p1 and phl p5; Bermuda grass proteinCyn d 5; Alternaria alternate proteins Alt a 1, Alt a 2 and Enolase (Alta 6); Birch protein Bet v1 and P14; German Cockroach proteins Bla g 1,Bla g 2, Bla g 3, Bla g 4, Bla g 5 and Bla g 6; Mugwort protein Art v 1;Russian thistle protein Sal k 1 and Sal k 2; peanut Ara h1, Ara h2, Arah3, Ara h4, Ara h5, Ara h6, plant profilins or lipid transfer proteinsor a human leukocyte antigen.Delivery Methods

Once formulated the compositions of the invention can be delivered to asubject in vivo using a variety of known routes and techniques. Forexample, a composition can be provided as an injectable solution,suspension or emulsion and administered via parenteral, subcutaneous,epidermal, intradermal, intramuscular, intraarterial, intraperitoneal,intravenous injection using a conventional needle and syringe, or usinga liquid jet injection system. Compositions can also be administeredtopically to skin or mucosal tissue, such as nasally, intratracheally,intestinal, rectally or vaginally, or provided as a finely divided spraysuitable for respiratory or pulmonary administration. Other modes ofadministration include oral administration, suppositories, sublingualadministration, and active or passive transdermal delivery techniques.

Where a peptide of the invention is to be administered, it is preferredto administer the peptide to a site in the body where it will have theability to contact suitable antigen presenting cells, and where it, orthey, will have the opportunity to contact T cells of the individual.Where an APC is to be administered, it is preferred to administer theAPC to a site in the body where it will have the ability to contact, andactivate, suitable T cells of the individual.

Delivery Regimes

Administration of the peptides/polynucleotides/cells (such as thecomposition containing a plurality of peptides) may be by any suitablemethod as described above. Suitable amounts of the peptide may bedetermined empirically, but typically are in the range given below. Asingle administration of each peptide may be sufficient to have abeneficial effect for the patient, but it will be appreciated that itmay be beneficial if the peptide is administered more than once, inwhich case typical administration regimes may be, for example, once ortwice a week for 2-4 weeks every 6 months, or once a day for a weekevery four to six months. As will be appreciated, each peptide orpolynucleotide, or combination of peptides and/or polynucleotides may beadministered to a patient singly or in combination.

Dosages for administration will depend upon a number of factorsincluding the nature of the composition, the route of administration andthe schedule and timing of the administration regime. Suitable doses ofa molecule of the invention may be in the order of up to 15 μg, up to 20μg, up to 25 μg, up to 30 μg, up to 50 μg, up to 100 μg, up to 500 μg ormore per administration. Suitable doses may be less than 15 μg, but atleast 1 ng, or at least 2 ng, or at least 5 ng, or at least 50 ng, orleast 100 ng, or at least 500 ng, or at least or at least 10μg. For somemolecules of the invention, the dose used may be higher, for example, upto 1 mg, up to 2 mg, up to 3 mg, up to 4 mg, up to 5 mg or higher. Suchdoses may be provided in a liquid formulation, at a concentrationsuitable to allow an appropriate volume for administration by theselected route.

Kits

The invention also relates to a combination of components describedherein suitable for use in a treatment of the invention which arepackaged in the form of a kit in a container. Such kits may comprise aseries of components to allow for a treatment of the invention. Forexample, a kit may comprise one or more different peptides,polynucleotides and/or cells of the invention, or one or more peptides,polynucleotides or cells of the invention and one or more additionaltherapeutic agents suitable for simultaneous administration, or forsequential or separate administration. The kit may optionally containother suitable reagent(s) or instructions and the like.

The invention is illustrated by the following Examples:

Example 1 MHC Class II Binding Search

The aim of this study is to identify a distinct panel of peptides withstrong affinities for the seven most common human MHC Class II HLA-DRB1*allotypes (covering in total around 63% of the allotypes found in theaverage Caucasian population). In order to identify binding peptides inthe House Dust Mite (HDM) allergens, Der p 1, Der p 2 and Der p 7, invitro binding assays have been performed on a subset of peptides fromthese allergenic proteins. Peptides for testing in the binding assayswere initially identified by an in silico approach known as “peptidethreading” (carried out by Biovation, Ltd., Aberdeen, Scotland, UK).This is a bioinformatic analysis of consecutive peptides from a sequencefor the potential to be accommodated within the binding groove of MHCclass II HLA-DR molecules. This subset of peptides was pre-screened forsolubility in an aqueous, acidic milieu and a final panel of 44 peptidesselected for testing in an in vitro MHC Class II binding assay.MethodsThe assay employed is a competitive MHC class II binding assay, whereineach peptide is analysed for its ability to displace a known controlbinder from each of the human MHC class II allotypes investigated. Theallotypes and control peptides used in this study are shown in Table 2:

TABLE 2 Control peptides used in the in vitro binding assays AllotypeControl Peptide Sequence DRB1*0101 Influenza haemagglutinin 307-319PKYVKQNTLKLAT (SEQ ID NO: 116) DRB1*0301Myco. tuberculosis/leprae hsp 65 2-16 AKTIAYDEEARRGLE (SEQ ID NO: 117)DRB1*0401 Influenza haemagglutinin 307-319 PKYVKQNTLKLAT(SEQ ID NO: 116) DRB1*0701 Influenza haemagglutinin 307-319PKYVKQNTLKLAT (SEQ ID NO: 116) DRB1*1101Influenza haemagglutinin 307-319 PKYVKQNTLKLAT (SEQ ID NO: 116)DRB1*1301 HLA-DQB1*0603 21-36 TERVRLVTRHIYNREE (SEQ ID NO: 118)DRB1*1501 Human myelin basic protein 85-99 ENPVVHFFKNIVTPR(SEQ ID NO: 119) DQB1*0602 Human Insulin B 1-15 FVNQHLCGSHLVEAL(SEQ ID NO: 120)Each of the 44 HDM peptides (which are shown in Tables 3A and 3B) wereanalysed in the competition assay and screened for relative bindingcompared to the control peptide. Due to the nature of the competitiveassay the data for each peptide is represented as a ratio of its ownIC50 to that of the control peptide. Thus, a peptide that has an IC50value that is parity to the control peptide has an identical bindingaffinity, while peptides with a ratio less than one have a higheraffinity and those with a ratio greater than one have a lower affinity.ResultsSolubility in aqueous solution is an essential criterion for a peptideto be an effective therapeutic agent. Therefore, as a consequence of thesolubility screen we will have eliminated very hydrophobic peptides witha high frequency of large hydrophobic amino acid residues in multiplebinding registers. This is a characteristic of promiscuous HLA-DRB1*binders. The data from the binding assays is shown in Table 3B. Therelative binding of each peptide is shown for each of the allotypes inthe study. The data shows that 24 of the 44 peptides tested bound to oneor more of the MHC Class II allotypes. A range of cross-reactivity isseen with 5 peptides binding only one allotype, 8 peptides binding two,9 peptides binding three and two peptides binding four different MHCClass II allotypes (red). It would also be expected that such peptideswould have the ability to bind similar allotypes that have not beentested through the homology of MHC structures. This can be seen in thecross-reactivity of peptides for DRB1*0101, *0401, *0701 and *1101 inseveral cases here. Also shown is the solubility status of the peptideat the highest concentrations in the aqueous solution of the bindingassay. The value illustrates the lowest concentration at which aninsoluble white precipitate is seen. There appears to be no significantnonspecific effect of the formation of precipitate in the assays.Several peptides that precipitate at high concentrations also bind toMHC class II; however, several also show no ability to compete with thecontrol peptides. It is to be expected that peptides liable to formprecipitates may exhibit high affinity and promiscuous binding due tothe presence of many hydrophobic residues.

The % purity of the peptides is indicated in Table 3A. This is ofsignificance as purities were seen to vary from 60-90%. This would havea considerable effect on the ability of a peptide to compete if it isrelatively impure. For example, HDM23A and HDM32 show low affinitybinding; however, they are of reduced purity (66.7% and 68.7%respectively) compared to other HDM peptides. Therefore, if purity istaken into consideration, they may in fact have an equivalent affinityto a peptide of a higher purity.

It can be seen that some MHC Class II allotypes bind to more peptidesthan others; this is probably to be expected as there is variabilitybetween the pocket positions in the different MHC class II bindinggrooves. There are however, also a number of well-characteriseddifferences between the affinities of the various control peptides.Clearly a high affinity control peptide will be more difficult todisplace by the competing HDM peptide resulting in the identification offewer binding peptides. This can be illustrated by the data presentedhere. For example, the Influenza Haemagglutinin 307-319 control peptide,has varying affinity according to the allotype, whereDRB1*0101>*0401>*0701>*1101. This is reflected in the number of bindersto each of the allotypes, where DRB1*0101 has the lowest number ofbinders (5) and DRB1*1101 has the highest (14). Furthermore, the bindingassay for DRB1*1501 is very stringent due to the high affinity of MyelinBasic Protein 85-99 for this allotype. In the high stringency screen theFel d 1 peptide EQVAQYKALPVVLENA (SEQ ID NO: 121), that was tested in anearlier study, gave a ratio of 0.97 indicating that high affinitybinders could be identified at this stringency.

In addition, to identify lower affinity binders, the assay was alsocarried out under less stringent conditions. All the Der p bindingpeptides were seen to have a high ratio when tested against thisallotype, showing they were low affinity binders compared to the controlpeptide. The DQA1*0102/DQB1*0602 binding assay uses a peptide from theB-chain of human insulin which is of lower affinity compared to thoseused in the DR assays. This dictates that the DQ assay is very sensitiveand tends to produce very low ratio values for the strongest binders tothis MHC Class II allotype. This sensitivity also accounts for therelatively higher number of DQ binding peptides within the panelscreened. Finally, on closer analysis, the peptides identified asligands for the DRB1*0101, *0401, *0701 superfamily, are found toincorporate a motif that is characteristic of promiscuous binders tothis family of allotypes where: P1=Y, F, W, L, I, V, or M (Largearomatic or hydrophobic residue), P6=S, T, C, A, P, V, I, M (small,non-charged residue) Out of the 16 peptides (e.g. HDM 21BRGKPFQLEAVFEANQNT (SEQ ID NO: 10)) identified as binders to all or acombination of these 3 allotypes, 14 (87.5%) contain this motif, whichsuggests that these are promiscuous binders with a range of affinitiesfor the 1-4-7 allotypes.

CONCLUSIONS

A range of peptides have been shown to have the capacity to bind the MHCClass II allotypes tested and can be tested for their ability tostimulate in vitro proliferation of CD4+ T cells and to stimulate T cellcytokine secretion.

TABLE 3A Residues in parent % Solubility Precipitation PeptideI Sequence molecule purity test in assay HDM01 IDLRQMRTVTPIR 112-12479.2 YES None (SEQ ID NO: 1) HDM02 RTVTPIRMQGGCG 118-130 79.6 YES None(SEQ ID NO: 2) HDM03C RNQSLDLAEQELVDCAS 149-167 60.1 YES NoneQH (SEQ ID NO: 3) HDM05 EYIQHNGVVQESY 179-191 77.5 YES None(SEQ ID NO: 122) HDM06 RYVAREQSCRRPN 193-205 79.7 YES None(SEQ ID NO: 4) HDM07 PNVNKIREALAQT 220-232 88.6 YES None (SEQ ID NO: 56)HDM08 NKIREALAQTHSA 223-235 87.6 YES None (SEQ ID NO: 123) HDM09AREALAQTHSAIAVI 226-239 69.6 YES 10000 μM (SEQ ID NO: 5) (2.9 mg/ml)HDM11 IGIKDLDAFRHYD 240-252 77.6 YES None (SEQ ID NO: 124) HDM12KDLDAFRHYDGRT 243-255 72.9 YES None (SEQ ID NO: 125) HDM13RTIIQRDNGYQPNY 254-267 70.7 NO None (SEQ ID NO: 126) HDM16ARNSWDTNWGDNGYG 287-300 70.00 YES None (SEQ ID NO: 127) HDM17NSVNVPSELDLRSLRT 105-120 74.5 YES None (SEQ ID NO: 128) HDM19DQVDVKDCANHEIKK 18-32 81.4 YES None (SEQ ID NO: 6) HDM20 CITHRGKPFQLEA44-56 77.4 YES None (SEQ ID NO: 7) HDM21 KPFQLEAVFEANQNT 50-64 88.7 YES2000 μM (SEQ ID NO: 8) (0.3 mg/ml) HDM21A KPFQLEAVFEANQNTK 50-65 90.10YES 50000 μM (SEQ ID NO: 9) (9.3 mg/ml) HDM21B RGKPFQLEAVFEANQNT 48-6482.60 YES 10000 μM (SEQ ID NO: 10) (1.98 mg/ml) HDM22A EAVFEANQNTKTAK55-68 90.30 YES None (SEQ ID NO: 11) HDM23A DGLEVDVPGIDPNACH 76-88 66.7YES None (SEQ ID NO: 12) HDM26A DGVLACAIATHAKIR 131-145 10000 μM(SEQ ID NO: 13) (1.5 mg/ml) HDM27 AKIEIKASLDGLE 67-79 65.9 YES 10000 μM(SEQ ID NO: 129) (1.4 mg/ml) HDM28 KAVDEAVAAIEKS 31-43 86.8 YES 10000 μM(SEQ ID NO: 130) (1.3 mg/ml) HDM29 ETFDPMKVPDHSD 44-56 84.7 YES None(SEQ ID NO: 131) HDM29A ETFDPMKVPDHSDK 44-57 91.7 YES None(SEQ ID NO: 132) HDM29B KSETFDPMKVPDHSD 42-56 92.5 YES 10000 μM(SEQ ID NO: 133) (1.7mg/ml) HDM30 DKFERHIGIIDLK 56-68 81.4 YES 50000 μM(SEQ ID NO: 14) (7.9 mg/ml) HDM31 IGIIDLKGELDMRN 62-75 10000 μM(SEQ ID NO: 134) (1.8 mg/ml) HDM31A HIGIIDLKGELDMRN 61-75 66.40 YES10000 μM (SEQ ID NO: 135) (1.7 mg/ml) HDM32 IDLKGELDMRNIQ 65-77 68.7 YES50000 μM (SEQ ID NO: 15) (7.7 mg/ml) HDM32A IDLKGELDMRNIQVR 65-79 85.20YES 50000 μM (SEQ ID NO: 136) (9.0 mg/ml) HDM33 LDMRNIQVRGLKQ 71-83 70.3YES None (SEQ ID NO: 16) HDM34 RNIQVRGLKQMKRVG 74-88 74.7 YES None(SEQ ID NO: 17) HDM35 RGLKQMKRVGDAN 79-91 84.00 YES None (SEQ ID NO: 18)HDM36 KRVGDANVKSEDG 85-97 82.9 YES None (SEQ ID NO: 137) HDM37ANVKSEDGVVKAH 90-102 76.5 YES None (SEQ ID NO: 138) HDM39 DDVVSMEYDLAYK109-121 84.9 NO* None (SEQ ID NO: 139) HDM39A HDDVVSMEYDLAYKL 108-12180.9 YES 10000 μM (SEQ ID NO: 19) (1.8 mg/ml) HDM40A VSMEYDLAYKLGDLH112-124 66.9 YES 10000 μM (SEQ ID NO: 20) (1.8 mg/ml) HDM48TAIFQDTVRAEMTK 187-200 79.1 YES 10000 μM (SEQ ID NO: 21) (1.6 mg/ml)HDM49 DTVRAEMTKVLAP 192-204 69.5 YES None (SEQ ID NO: 22) HDM50KVLAPAFKKELER 200-212 90.8 YES None (SEQ ID NO: 140) HDM51 VDFKGELAMRNIE65-77 79.8 YES 10000 μM (SEQ ID NO: 23) (1.5 mg/ml) HDM51AVDFKGELAMRNIEAR 65-79 82.1 YES None (SEQ ID NO: 24)

TABLE 3B DQA1*0102 Peptide DRB1*0101 DRB1*0301 DRB1*0401 DRB1*0701DRB1*1101 DRB1*1301 DRB1*1501 QB1*0602 HDM01 19.23 16 HDM02 80 0.03HDM03C 0.16 HDM05 HDM06 30.36 0.86 HDM07 HDM08 HDM09A 0.49 21.15 200HDM11 HDM12 HDM13 HDM16A HDM17 HDM19 HDM20 1.1 28 242.11 2.37 HDM21 9211.15 11.73 HDM21A 200 52.17 10.27 HDM21B 13.5 0.78 4.1 HDM22A 328.6 80HDM23A 347 0.76 HDM26A 42.3 16.28 0.61 HDM27 HDM28 HDM29 HDM29A HDM29BHDM30 6.2 HDM31 HDM31A HDM32A HDM33 46.51 41.5 263.16 HDM34 3.38 3.7769.23 HDM35 1.26 HDM36 HDM37 HDM39 HDM39A 76.19 0.71 0.1 HDM40A 2.29 6HDM48 211.26 15.71 13.57 HDM49 671.43 1.7 HDM50 HDM51 20.93 30.91

Example 2

Homology Search

The sequences of each of the 24 peptides identified above as MHC ClassII-binding were used to probe the sequence of the alternative protein inthe dust mite allergen group from which the parent sequence derived. Forexample, peptide HDM01 in Table 3A is from Der p 1, therefore thesequence of HDM01 was used to probe for a homologous sequence in Derf 1. The same practice was applied for all 24 peptides identified above.The peptides identified in Example 1 and Example 2 are shown in Tables 4to 6.

TABLE 4 Residues SEQ  Peptide in Parent in ID Table 3A/B moleculeSequence parent NO: HDM01 Der p 1 IDLRQMRTVTPIR 112-124 1 Der f 1LDLRSLRTVTPIR 113-125 25 HDM02 Der p 1 RTVTPIRMQGGCG 118-130 2 Der f 1RTVTPIRMQGGCG 119-131 2 HDM03C Der p 1 RNQSLDLAEQELVDCASQH 149-167 3Der f 1 RNTSLDLSEQELVDCASQH 150-168 27 HDM06 Der p 1 RYVAREQSCRRPN193-205 4 Der f 1 PYVAREQRCRRPN 194-206 28 HDM09A Der p 1 REALAQTHSAIAVI226-239 5 Der f 1 REALTOTHTAIAVI 227-240 29

TABLE 5 Residues SEQ Peptide in Parent in ID Table 3A/B moleculeSequence parent NO: HDM19 Der p 2 DQVDVKDCANHEIKK 18-32 6 Der f 2DQVDVKDCANNEIKK 18-32 30 HDM20 Der p 2 CIIHRGKPFQLEA 44-56 7 Der f 2CIIHRGKPFTLEA 44-56 31 HDM21 Der p 2 KPFQLEAVFEANQNT 50-64 8 Der f 2KPFTLEALFDANQATT 50-64 32 HDM21A Der p 2 KPFQLEAVFEANQNTK 50-65 9Der f 2 KPFTLEALFDANQATTK 50-65 33 HDM21B Der p 2 RGKPFQLEAVFEANQNT48-64 10 Der f 2 RGKPFTLEALFDANQATT 48-64 34 HDM22A Der p 2EAVFEANQNTKTAK 55-68 11 Der f 2 EALFDANONTKTAK 55-68 35 HDM23A Der p 2DGLEVDVPGIDPNACH 76-88 12 Der f 2 DGLEIDVPGIDTNACH 76-88 36 HDM26ADer p 2 DGVLACAIATHAKIR 131-145 13 Der f 2 NGVLACAIATHGKIR 131-145 37

TABLE 6 Residues SEQ  Peptide in Parent in ID Table 3A/B moleculeSequence parent NO: HDM30 Der p 7 DKFERHIGIIDLK 56-68 14 Der f 7DKFERHVGIVDFK 56-68 38 HDM32 Der p 7 IDLKGELDMRNIQ 65-77 15 Der f 7VDFKGELAMRATIE 65-77 39 HDM33 Der p 7 LDMRNIQVRGLKQ 71-83 16 Der f 7LAMRNTEARGLKQ 71-83 40 HDM34 Der p 7 RNIQVRGLKQMKRVG 74-88 17 Der f 7RNTEARGLKOMKRQG 74-88 41 HDM35 Der p 7 RGLKQMKRVGDAN 79-91 18 Der f 7RGLKOIKRCGDAN 79-91 42 HDM39A Der p 7 HDDVVSMEYDLAYKL 108-122 19 Der f 7HDDIVSMEYDLAYWL 108-122 43 HDM40A Der p 7 VSMEYDLAYKLGDLH 112-126 20Der f 7 VSMEYDLAYKLGDLH 112-126 20 HDM48 Der p 7 TAIFQDTVRAEMTK 187-20021 Der f 7 TAIFCDTVRKEMTK 187-200 45 HDM49 Der p 7 DTVRAEMTKVLAP 192-20422 Der f 7 DTVRKEMTKVLAP 192-204 46 HDM51 Der f 7 VDFKGELAMRNIE 65-77 23Der p 7 IDLKGELDMRATIQ 65-77 15 HDM51A Der f 7 VDFKGELAMRNIEAR 65-79 24Der p 7 IDLKGELDMRATIQVR 65-79 47In Table 4, the sequence of Der p 1 from which the “residues in parent”positions are derived is the publically available sequence with NCBIAccession No. P08176. The corresponding sequences for Der p 2 (Table 5)and Der p 7 (Table 6) are NCBI Accession Nos. P49278 and P49273,respectively. The sequence for Der f 1 is taken from NCBI Accession No.P16311, Der f 2 is from NCBI Accession No. Q00855 and Der f 7 is fromNCBI Accession No. Q26456.

Example 3

MHC Class II Binding Search

The aim of this study is to identify a distinct panel of peptides withstrong affinities for the seven most common human MHC Class II HLA-DRB1*allotypes (covering in total around 63% of the allotypes found in theaverage Caucasian population). In order to identify binding peptides inthe major House dust mite allergens Der p 1, Der p 2 and Der p 7.Peptides were identified by an in silico approach known as “peptidethreading” using the commercially available EpiMatrix algorithm (EpiVaxInc.) This is a bioinformatic analysis of peptides from a sequence forthe potential to be accommodated within the binding groove of MHC classII HLA-DR molecules. EpiMatrix is a matrix-based algorithm that ranks 10amino acid long segments, overlapping by 9 amino acids, from anypolypeptide sequence by estimated probability of binding to each of theselected MHC molecules. (De Groot et al., AIDS Research and HumanRetroviruses 13:539-41 (1997)). The procedure for developing matrixmotifs was published by Schafer et al, 16 Vaccine 1998 (1998). In thisExample, binding potential for HLA DR1, DR2, DR3, DR4, DR7, DR8, DR11,DR13 and DR15 is assessed. Putative MHC ligands are selected by scoringeach 10-mer frame in a protein sequence. This score is derived bycomparing the sequence of the 10-mer to the matrix of 10 amino acidsequences known to bind to each MHC allele. Retrospective studies havedemonstrated that EpiMatrix accurately predicts published MHC ligands(Jesdale et al., in Vaccines '97 (Cold Spring Harbor Press, Cold SpringHarbor, N.Y., 1997)). Successful Prediction of peptides which bind tomultiple MHC molecules has also been confirmed.

Estimated probability of binding to a selected MHC molecule iscalculated by EpiMatrix as follows. The peptides are scored byestimating the relative promotion or inhibition of binding for eachamino acid, compared to known MHC binders for a given MHC allele. Thisinformation is summed across the peptide and a summary score (EMX score)is assigned to the entire peptide. After comparing the EMX score to thescores of known MHC ligands, EpiMatrix arrives at an “estimated bindingprobability” (abbreviated as EBP, but not strictly a probability). TheEBP describes the proportion of peptides with EpiMatrix scores as highor higher that will bind to a given MHC molecule. EBPs range from 100%(highly likely to bind) to less than 1% (very unlikely to bind).

EpiMatrix analyses were performed on the entire sequence of the Der p 1as published in the NCBI database (NCBI accession no: P08176). Thisanalysis identified core peptides (and their flanking sequences) derivedfrom the above sequences which are predicted to have good MHC class-IIbinding. These sequences are shown below in Table 7A. Tables 7B and 7Cshow the sequences for the equivalent analyses of known sequences of Derp 2 and Der p 7, respectively (NCBI accession nos. P49278 and P49273).

In Tables 7A-C: “Residues in sequence” gives the location of the peptidewithin the sequences that were analysed. The core peptide (middle aminoacids in bold) defines the actual binding sequence that was identifiedduring the analysis. The stabilizing flanks (N-terminal and C-terminal,not bold) were included for use with the core sequence and are typicallyrequired to aid manufacture of the peptides. “Number of hits” refers tothe number of high predicted binding affinities for all MHC types testedwithin the sequence. The “EpiMatrix Cluster Score” is derived from thenumber of hits normalized for the length of the cluster. Cluster Scoreis thus the excess or shortfall in predicted aggregate MHC bindingproperties relative to a random peptide standard. A score of 10 or aboveis considered to indicate broad MHC binding properties. Epivax alsoanalysed hydrophobicity of peptides containing epitopes. Scores ofgreater than 1 are considered to be unsuitable for administration and/ormanufacture.

TABLE 7A Der p 1 EpiMatrix RESIDUES IN EpiMatrix CLUSTER INPUT SEQUENCEHITS SCORE SEQ SEQUENCE (Incl. Hydro- (Excl (Excl ID (NCBI no.) FLANKS)SEQUENCE phobicity FLANKS) FLANKS) NO: P08176 1-21 MKIVLAIASLL 1.42 2238.91 105 ALSAVYARPS P08176 51-67 LESVKYVQSNG −0.15 6 10.87 106 GAINHLP08176 72-88 LDEFKNRFLMS −0.49 6 10.55 107 AEAFEH P08176 111-134EIDLRQMRTVTP −0.24 16 26.34 108 IRMQGGCGSCWA P08176 142-159 ESAYLAYRNQSL−0.91 10 16.43 109 DLAEQE P08176 188-209 QESYYRYVAREQS −1.70 14 24.92110 CRRPNAQRF P08176 296-313 DNGYGYFAANID −0.08 7 10.24 86 LMMIEE

TABLE 7B Der p 2 EpiMatrix RESIDUES IN EpiMatrix CLUSTER INPUT SEQUENCEHITS SCORE SEQ SEQUENCE (Incl. Hydro- (Excl (Excl ID (NCBI no.) FLANKS)SEQUENCE phobicity FLANKS) FLANKS) NO: P49278 1-22 MMYKILCLSLLVA 1.24 1421.8 112 AVARDQVDV P49278 42-63 EPCIIHRGKPFQL -0.50 10 14.62 113EAVFEANQN

TABLE 7C Der p 7 EpiMatrix RESIDUES IN EpiMatrix CLUSTER INPUT SEQUENCEHITS SCORE SEQ SEQUENCE (Incl. Hydro- (Excl (Excl ID (NCBI no.) FLANKS)SEQUENCE phobicity FLANKS) FLANKS) NO: P49273 1-17 MMKLLLIAAAA 2.2 1220.16 114 FVAVSA P49273 70-92 ELDMRNIQVRGLK -0.71 9 12.3 115 QMKRVGDANV

Example 4

Selection of Peptides for Further Testing

Based on the peptides and epitopes identified in Examples 1 to 3, theinventors selected the peptides shown in Table 8 for further testing.Some of the peptides selected can be considered to be variants of thepeptides of Example 1 to 3, but are also considered to be peptides ofthe invention. In particular, residues in bold in Table 8 indicatealterations from the corresponding residue in the native sequence of theparent protein. These alterations reduce the formation of peptide dimersand improve solubility without diminishing the functionality of apeptide as a T cell epitope. The alterations shown are the replacementof a cysteine (C) in the native sequence with a serine (S) or5-aminobutyric acid (

), or cystine (Ĉ) as indicated.

Additionally, some sequences may comprise more or fewer of the residuesof the parent protein from which they derive, when compared to thesequences of the peptides of Examples 1 to 3. Thus, such sequences canbe considered to represent truncation or extension variants of thepeptides of Examples 1 to 3. For example, Peptide HDM03F corresponds toresides 149-165 of Der p1. HDM03E corresponds to residues 149-167.Accordingly, HDM03F can be considered to be a truncation variant ofHDM03E formed by removal of 2 residues from the N terminus of HDM03E.The “residues in parent” positions in Table 8 refer to the sequences ofDer p 1, Der p 2 and Der p 7 as used in Tables 4 to 7. Those peptidesindicated in Table 8 which have an N terminal glutamate (E) residue, forexample HDM03K, L, V and W, may have the glutamate replaced withpyroglutamate to improve stability during manufacture, without affectingfunction of the peptide. The data from further testing of these peptides(Example 5) is typically obtained using peptides where such replacementhas taken place.

TABLE 8 Residues SEQ Parent in ID Peptide molecule Sequence parent NO:HDM01 Der p 1 IDLRQMRTVTPIR 112-124 1 HDM01A Der p 1 IDLRQMRTVTPIRMQGGSG112-130 48 HDM02A Der p 1 RTVTPIRMQGGSG 118-130 49 HDM02B Der p 1RTVTPIRMQGG

G 118-130 50 HDM03D Der p 1 RNQSLDLAEQELVDSASQH 149-167 51 HDM03EDer p 1 RNQSLDLAEQELVD

ASQH 149-167 52 HDM03F Der p 1 RNQSLDLAEQELVDSAS 149-165 53 HDM03GDer p 1 QSLDLAEQELVD

ASQHG 151-168 89 HDM03H Der p 1 LDLAEQELVD

ASQHG 153-168 90 HDM03J Der p 1 LAEQELVD

ASQHG 155-168 91 HDM03K Der p 1 EQELVD

ASQHG 157-168 92 HDM03L Der p 1 ELVD

ASQHG 159-168 93 HDM03M Der p 1 RNQSLDLAEQELVDCASQHG 149-168 94 HDM03NDer p 1 RNQSLDLAEQELVD{circumflex over (C)}ASQHG 149-168 95 HDM03PDer p 1 SAYLAHRNQSLDLAEQELVDCAS 143-166 96 HDM03R Der p 1QSLDLAEQELVDSASQHG 151-168 97 HDM03S Der p 1 LDLAEQELVDSASQHG 153-168 98HDM03T Der p 1 LAEQELVDSASQHG 155-168 99 HDM03V Der p 1 EQELVDSASQHG157-168 100 HDM03W Der p 1 ELVDSASQHG 159-168 101 HDM06A Der p 1RYVAREQSSRRP 193-205 54 HDM06B Der p 1 RYVAREQS

RRP 193-205 55 HDM07 Der p 1 PNVNKIREALAQT 220-232 56 HDM09A Der p 1REALAQTHSAIAVI 226-239 5 HDM19A Der p 2 DQVDVKDSANHEIKK 18-32 57 HDM19BDer p 2 DQVDVKD

ANHEIKK 18-32 58 HDM20A Der p 2 IIHRGKPFQLEA 45-56 59 HDM20B Der p 2SIIHRGKPFQLEA 44-56 60 HDM21 Der p 2 KPFQLEAVFEANQNT 50-64 8 HDM21ADer p 2 KPFQLEAVFEANQNTK 50-65 9 HDM21B Der p 2 RGKPFQLEAVFEANQNT 48-6410 HDM22A Der p 2 EAVFEANQNTKTAK 55-68 11 HDM23B Der p 2 GLEVDVPGIDPNA77-86 61 HDM23C Der p 2 GLEVDVPGIDPNASH 77-88 62 HDM26B Der p 2GVLASAIATHAKIR 132-145 63 HDM26C Der p 2 GVLA

AIATHAKIR 132-145 64 HDM30 Der p 7 DKFERHIGIIDLK 56-68 14 HDM32 Der p 7IDLKGELDMRNIQ 65-77 15 HDM33 Der p 7 LDMRNIQVRGLKQ 71-83 16 HDM34Der p 7 RNIQVRGLKQMKRVG 74-88 17 HDM35A Der p 7 RGLKQMKRVGDANV 79-92 65HDM39A Der p 7 HDDVVSMEYDLAYKL 108-121 19 HDM39B Der p 7HDDVVSMEYDLAYKLGDLH 108-125 66 HDM40A Der p 7 VSMEYDLAYKLGDLH 112-124 20HDM40B Der p 7 VSMEYDLAYKLGDL 112-123 67 HDM48 Der p 7 TAIFQDTVRAEMTK187-200 21 HDM48A Der p 7 TAIFQDTVRAEMTKVLAP 187-204 68 HDM49 Der p 7DTVRAEMTKVLAP 192-204 22 HDM51 Der p 7 VDFKGELAMRNIE 65-77 23 HDM51ADer p 7 VDFKGELAMRNIEAR 65-79 24 HDM100 Der p 1 RFGISNYCQIYPPNVNK208-224 69 HDM100A Der p 1 RFGISNYSQIYPPNVNK 208-224 70 HDM100B Der p 1RFGISNY

QIYPPNVNK 208-224 71 HDM101 Der p 1 NYCQIYPPNVNKIREA 213-228 72 HDM101ADer p 1 NYSQIYPPNVNKIREA 213-228 73 HDM101B Der p 1 NY

QIYPPNVNKIREA 213-228 74 HDM102 Der p 1 NAQRFGISNYCQI 205-217 75 HDM102ADer p 1 NAQRFGISNYSQI 205-217 76 HDM102B Der p 1 NAQRFGISNY

QI 205-217 77 HDM103 Der p 2 KGQQYDIKYTWNVPKIAP 99-116 78 HDM104 Der p 2WNVPKIAPKSENV 109-121 79 HDM201 Der p 1 ESVKYVQSNGGAI 52-64 80 HDM202Der p 1 DEFKNRFLMSAEAFE 73-87 81 HDM202D Der p 1 FKNRFLMSAEA 75-85 102HDM202E Der p 1 FKNRFLMSAE 75-84 103 HDM202H Der p 1 EFKNRFLMSAE 74-84104 HDM203A Der p 1 DLRQMRTVTPIRMQGGCGS 113-131 82 HDM203B Der p 1DLRQMRTVTPIRMQGGSGS 113-131 83 HDM204 Der p 1 SAYLAYRNQSLDLA 143-156 84HDM205 Der p 1 SYYRYVAREQS 190-199 85 HDM206 Der p 1 DNGYGYFAANIDLMMIEE296-313 86 HDM206A Der p 1 NGYGYFAANIDLMM 297-310 87 HDM207 Der p 7DMRNIQVRGLKQMKRVGD 72-104 88

Example 5

Cytokine Release Assay and Selection of Peptide Combinations

Cytokine secretion profiles from PBMC's are analysed in response to thepeptide stimulation using the peptides from Example 3. Supernatants fromthe cytokine release assay are tested for the presence of 2 cytokines,IFN-γ and IL-13, using ELISA assays. Cytokine secretion profiles fromPBMC's were analysed in response to the peptide stimulation using thepeptides indicated. Supernatants from the cytokine release assay weretested for the presence of 2 cytokines, IFN-γ and IL-13, using either anELISA assay or a multiplex bead array assay.

A typical cytokine release assay requires 40×10⁶ PBMC's per subject. Inmore detail, 250 μl of a 200 μg/ml solution of the appropriate antigenor peptide concentration is distributed into the appropriate wells of 48well plates. Plates are the incubated in a humidified 5% CO₂ incubatorat 37° C. for a maximum of 4 hours. 250 μl of a 5×10⁶ cell/ml PBMCsuspension is then added to each well and the plates returned to theincubator for 5 days. Following stimulation, samples of culturesupernatant are harvested for testing by ELISA or multiplex bead assayaccording to standard protocols.

Il-13 and IFN-gamma responses to each peptide were scored as positive Tcell epitopes provided the amount of cytokine produced in the well forthat peptide exceeded 100 pg/ml, i.e. 100 pg per 1.25×10⁶ cells. Thus,an individual was considered to have responded to a peptide if cellsfrom that individual yielded a response greater than 100 pg/ml foreither 11-13 or IFN-gamma. The percentage of responders to each peptideis shown in FIG. 2.

The top five peptides by percentage of individuals with an 11-13 orIFN-gamma response greater than 100 pg/ml are HDM203B, HDM201, HDM205,HDM203A and HDM202, and (SEQ ID NOS. 83, 80, 85, 82 and 81).

HDM203A and 203B are variants of the same sequence with 203B modifiedsuch that a serine replaces a cysteine (at the third residue from the Cterminus) to achieve better manufacturability and stability. Thus apreferred combination of peptides should comprise at least one of thesepeptides or a variant thereof.

The next most potent peptides are HDM09A, HDM03D, HDM03E, HDM101,HDM101A, HDM101B (SEQ ID NOS: 5, 51, 52, 72, 73 and 74). A preferredpeptide combination may typically comprise at least one additionalpeptide selected from this group. Of this group HDM03D and HDM03E aresequence variants of each other with serine and aminobutyric acid(respectively) replacing cysteine (at the fifth residue from the Cterminus of the native sequence of Der p 1) to achieve bettermanufacturability and stability. These sequences are consideredequivalent.

Further variants of HDM03, namely HDM03V and HDM03W (SEQ ID NO. 100 and101) are also considered to be suitable. These variants are fragmentscomprising a truncation down to the last eleven or ten (respectively) Cterminal residues of HDM03D. These peptides are not included in theassay described above, but on testing are considered to be at leastequivalent to HDM03D (data not shown).

HDM101, HDM101A, and HDM101B are also sequence variants of each other,with HDM101A having a serine and HDM 101B having an aminobutyric acidreplacing a cysteine in HDM101 (third residue from the N terminus). Allthree HDM101 series peptides are considered equivalent, with HDM101A orHDM101B preferred for manufacturability and stability.

Of the remainder peptides tested the following have responses in >25% ofthe individuals tested: HDM01 [Der p1], HDM01A [Der p1], HDM06A [Derp2], HDM07 [Der p1], HDM19A [Der p2], HDM21A [Der p2], HDM23C [Der p2],HDM26B [Der p2], HDM35A [Der p7], HDM48 [Der p7], HDM51A [Der f 7],HDM102A [Der p1], HDM204 [Der p1] and HDM206 [Der p1] (SEQ ID NOS. 1,48, 54, 56, 57, 9, 62, 63, 65, 21, 24, 76, 84, and 86 respectively). Apreferred peptide combination may typically comprise at least oneadditional peptide selected from this group. When considering whichadditional peptides to add to the mixture, representatives from thisfinal group should preferably be chosen from epitopes drawn from Der p2and Der p7 since the previous groups are dominated by Der p 1. HDM26B[Der p2] and HDM 35A [Der p′7] are particularly preferred. Additionalstudies (data not shown) demonstrate that these are the best performingpeptides from Der p 2 and Der p 7 respectively.

FIG. 3 shows the number of individuals who respond to a core mixture ofHDM201, HDM202, HDM203B and HDM205. The incremental effect of addingHDM03D and HDM101A, and the further incremental effect of adding HDM26Band HDM35A is also shown. The benefit of adding epitopes from the secondand third group of peptides is clearly shown.

Importantly, adding peptides 03D,26B,35A,101A to the core mixtureconverted 4 individuals from non-responders to responders. It is alsoapparent that removing one of the peptides 201, 202, 203B or 205 fromthe mixture would not reduce the number of overall responders to theproposed mixtures as most people have three or four responses to thispeptide group. This is demonstrated in FIG. 4, which shows similarresults for a core mixture of HDM201, HDM203B and HDM205.

The invention claimed is:
 1. A composition comprising at least onepharmaceutically acceptable diluent or carrier and-a polypeptideconsisting of the sequence of SEQ ID NO:
 65. 2. The compositionaccording to claim 1, which is a solution in which said polypeptide ispresent at a concentration in the range of 0.03 to 200 nmol/ml, 0.3 to200 nmol/ml or 30 to 120 nmol/ml.
 3. The composition according to claim1, which is formulated for oral administration, nasal administration,epicutaneous administration, subcutaneous administration, sublingualadministration, intradermal administration, buccal administration or foradministration by inhalation or by injection.